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• Chennai, Bangalore & Online: 93450 45466Coimbatore: 95978 88270Madurai: 97900 94102Pondicherry: 93635 21112 • SQL Interview Questions and Answers SQL Interview Questions and Answers The SQL database language is a powerful database language that can be used in a variety of ways to manage data. SQL is the most commonly used database management system in the world. It is used by companies of all sizes, from small businesses to large corporations. With SQL, you can easily manage and query data. The FITA Academy offers this Introduction to SQL Course in Chennai, which will give you the skills you need for a successful SQL interview. It is a language that can be used in a variety of different applications. If you’re considering a career in database development, or if you’re just looking to brush up on your skills, these SQL interview questions and answers from FITA Academy are a great place to start. A database is a collection of information organised into records within tables. For example, there might be customers, orders, inventory, etc. Each record represents something about one customer, order, item, etc. SQL stands for Structured Query Language. It is a computer programming language. The main purpose of SQL is to help people access, modify, organize, store, and retrieve data in a relational database. In general, data refers to any information stored in a computer. This includes text files, spreadsheets, databases, web pages, and so on. The stored information is called “data”. SQL was designed to make it easier to work with data. Before SQL came along, programmers had to create their own programs to manipulate data. There were no standard commands or procedures like in SQL. Nowadays, everything from personal computers to huge supercomputers have SQL installed as part of their operating systems. SQL is similar to Java and C# but not quite the same. Unlike other languages, SQL has its own syntax and keywords. You won’t find anything like this in C++ or Java. There are many types of databases, and these are the major ones. • Relational Database • Object-Oriented Database • Distributed Database • NoSQL Database • Graph Database • Cloud Database • Centralization Database • Operational Database A relational database stores information in rows and columns. For example, a person’s name might appear in the first row, address in the second row, phone number in the third row, and salary in the fourth row. These four items are combined together, making them one row. Rows are added together to form a table. Tables are then joined together to form a database. An object-oriented database is very similar to a relational database except that instead of storing data in tables, objects (also known as classes) are created. Objects are collections of properties. Properties are attributes that define what each object looks like. When you add more than one property to an object, they become linked, forming a hierarchy. A distributed database is a type of database where data is split across multiple servers. Servers may be located anywhere around the world. By having all of the data stored at many different locations, you get more performance out of your system. NoSQL stands for non-relational. It means that the data doesn’t need to be organized into tables or rows. Instead, data can be structured differently. This makes it easier to access and process data quickly. Graph databases organise information based on relationships between things. They follow directions such as a parent, child, sibling, friend, employee, etc. This allows users to navigate through the data easily. Cloud Database is a type of software used to store large amounts of data. This data is accessed over the internet by using a web browser. Examples include Google Drive, DropBox, Microsoft SkyDrive, Amazon S3, etc. The operational database is a database that is used for tracking data throughout the day. The data isn’t changed, just updated. If something goes wrong, you can use the operational database to track down problems. DBMS stands for database management system. It manages the storage and retrieval of data within a computer. Some examples are Oracle, MySQL, PostgreSQL, MSSQL, etc. RDBMS stands for relational database management systems. It is also called a traditional database management system. It stores data in tables. It uses SQL queries to retrieve data. Database Management Systems(DBMSs) are applications that manage data. RDBMSs are designed specifically for managing data. Tableau Software is a popular tool for visualizing data. Data is displayed with shapes and colors. You can create charts, graphs, maps, and dashboards using this tool. MySQL is a free open-source database platform. It is built on top of the UNIX operating system. It has easy-to-learn commands and syntax. It is also fast and reliable. If you’re looking to land a MySQL position in the near future, then you’ll want to take advantage of FITA Academy’s online MySQL Online Course. This program will help you learn all the important basics for a successful interview, including how to use SQL, navigate databases, and write effective queries. If you learn these things, you’ll be well on your way to landing the job of your dreams. SQL stands for Structured Query Language. It is a standard programming language used to query databases. MySQL is a programmable database engine. It is written in C++. MySQL is faster than SQL because it does not require parsing. The Four Subsets of SQL are the following: • Data definition language (DDL) • Data manipulation language (DML) • Data control language (DCL) • Transaction Control Language (TCL) Data Definition Language (DDL) is used to create, alter, and drop objects in a database. DDL is also known as CREATE, ALTER, DROP statements. These statements help us define the structure of our database. Data Manipulation Language (Dml) helps us change the data stored in our database. We may want to add or delete records. We may need to insert new rows into existing tables. Data Control Language (DCL is used to set up permissions for accessing various parts of the database. There are two types of DCL. One is user-based access control, and other is table-based access control. Transaction Control Language (TCL), is used to start, commit, rollback transactions. In plain English, we can say TCL is used to ensure the consistency of database. Tables are collection of related data. Fields store information about each record stored in a table. Let’s take the below given example. Here there is a table named “student“. Its field name is “name”. This table contains three records as shown below. Here first row shows the student’s name, second row shows his address, and third row shows his phone number. Table Name: `STUDENT` FIELDS: \| Name \| Address \| Phone Number FITA Academy offers interactive sessions that cover the most common sql interview questions and answers asked in SQL interviews. We also provide practice exams that help students assess their skills and weaknesses. We offers a variety of courses, from beginner to advanced, to suit everyone’s needs. • The constraints are mandatory. • They have no effect when the database is created. • They do not affect the size of the database. • They protect against errors. • They enforce referential integrity. A constraint is enforced at the time of creation of the Database. To explain you in better way imagine any relationship like mother father son daughter etc.. all these relationships should be present only once in the parent table so that child table will get associated with one and only one parent table. For eg. if i have a customer table then it can associate many different supplier table based on the type of product ordered by customer. In this case the reference from both sides must be unique and hence constraint comes here. If the same relation exists twice than it will lead to orphan record. Hence we make sure that this relation exist only once so that it will always get associated with one and every time. It is nothing but a unique attribute in the database. It becomes primary because it uniquely identifies a single record. A unique value is required so that we can identify any particular record in the database without ambiguity. When we create a relationship between two tables, then we need a column which holds the identity of one table. So that we know what record it needs to match, then those identities become known as foreign key columns. Unique keys are also called primary keys. They are primary because they’re used to uniquely identify a table. They are also key because they act as a link between two otherwise unrelated rows or attributes. They are unique because they contain values that don’t repeat across distinct rows. Foreign keys are columns that link rows of two tables. Let’s say I have a student table (“Students”), and a table (“Student_grades”). Every student has a unique ID number. We could use that number as a foreign key in “Student_grade” to connect their grades directly to them. Students who have higher grade point averages receive higher numbers. Normalizing means normalizing data into smaller units. There is no exact definition for normalization. However, it usually involves: • Reducing the number of fields within rows. If you notice, most relational databases contain fewer fields per row than non-relational databases. • Reducing redundancy by combining related items into fewer rows. • Relieving yourself of having to look up multiple tables to find the right combination of data. The goal is to remove redundant and unnecessary data, reducing the chance of storing incorrect or extra data. In general normalized data is easier to query, update, and maintain. One of the uses of foreign keys is to ensure that each unique entry in a foreign table corresponds to an entry in the primary table. This ensures that the structure of the foreign table matches the structure of its corresponding primary table. The other use of foreign keys is to enforce referential integrity. Some database systems allow referential integrity to be violated by deleting records from the secondary table and adding new records to the primary table instead. Inconsistent dependencies occur when one piece of code depends on a set of objects that may not exist. Denormalization is a form of data reduction in which data is duplicated. This includes all forms of duplication, such as repeating data (e.g., a table may contain three copies of the telephone number with different area codes) or repeating groups of data (a customer name can be repeated many times). In this case, denormalization means that duplicate data is stored in fewer places rather than more. Arithmetic and logical operations are performed with operators. All databases support the same basic types of arithmetic operators. These include addition (+), subtraction (-), multiplication (), division (/), exponentiation (), modulus (%) and bitwise AND (&), OR (\|), XOR (~) and NOT (!). Logical operators like AND, OR and NOT are often referred to as Boolean operators. SQL operates are: • Arithmetic operators • Logical operators • Comparison operators • Bitwise operators • Compound operators • String operators An index is a special type of data structure designed to speed up certain kinds of queries. It stores information about your data so that when you run a query against it, you don’t need to scan every record. A unique index is an index where no two values for the column being indexed can have the same value. Example: if you had a table containing employee names and phone numbers, you might want to create a unique index on the first name column to make sure that only one person has the exact same name. We use indexes because they provide several benefits, including faster retrieval and insertion, improved concurrency, and better space utilization. Clustered index refers to a kind of database indexing technique. In general, a clustered index is always at the end of the B-tree. That’s why it is called “clustered”. Non-Clustered indices are usually created on columns that are part of the PRIMARY KEY. They’re also useful when you need to find rows based on some other criteria besides their key. Joins are used to combine sets of related data together into a single result set. The most common join operation is the INNER JOIN, which combines the results of two tables according to matching conditions specified using WHERE clauses. When both tables have the same structure, there are four possible ways to write a simple SELECT statement. The simplest way is to list them in order: Inner Join is a SQL operator. It allows us to get records from two tables that share a relationship between each other. The output will show all the records from the first table along with any matching records from the second table. If there is no match, then the corresponding field will be null. Self-join is a type of join in relational databases. This means that the result table includes all the records from the original table as well as matching records from the same table itself. In a cross join, multiple tables are combined into one result set. The final result set should include all combinations of the individual elements. Outer Join is a SQL operator that combines rows from two tables into a single result set by linking them through a third table. The outer joined table contains all the rows from the first table, even those that do not have matches in the second table. Left or right outer join is similar to an outer join, but instead of returning all unmatched rows from the first table, it returns only those rows from the first table where the condition does not match. Right outer join is similar to left outer join but instead of matching the condition in the middle table it looks for mismatches in the second table. Although no rows are found in the second table, the first table is returned as a result. This can occur when the first row in the second table has no matches in the first table. Full outer join is like left or right Join except that it includes all matched rows regardless of whether they come from the left or right table. By default, this is true for left joins, but for right joins, it defaults to false. Our online course will help you learn the basics of SQL. This will help you prepare for a successful SQL interview whether you’re new to SQL or just need a refresher, our course is designed to help you succeed. Start learning how to ace your next SQL interview today! Nested query is a subquery where the result of the nested query becomes part of the outer query. This helps us perform complex queries without using loops. In the world of data management, views are virtual tables that emulate specific tables. Despite providing easy access to large amounts of data, they cannot stand alone. Because views are essentially just queries saved in text files, they tend to run very slowly. On the other hand, tables are physically organized objects that store information. They’re often much faster than views because they don’t require text encoding and decoding. • SELECT – Retrieves rows that meet certain criteria. • INSERT – Inserts new rows into a database table. • UPDATE – Changes existing values in a table. Set operators are used to find common elements between two sets. They can be used on any column in your table. The four different types of set operators are: • Union (U) – Find all records that exist in both collections. • Intersection (INTERSECT) – Finding all records that exist in at least one collection. • Except (EXCEPT) – Only include items that aren’t included in either list. • Symmetric Difference (SYMMETRIC DIFFERENCE) – Find all records that exist only in one collection but not the other. • IN – Finds records which belong within a given range. • BETWEEN – Finds records having specified date or time range. Where Clause shows conditions on particular columns whereas Having clauses show conditions among particular groups of result set. It stands for Atomicity, Consistency, Isolation, Durability. It means that all changes made by transactions should be valid and remain visible throughout the system. A Cursor is an object in SQL Server Management Studio that allows you to view data. You use cursors to browse through result sets produced by complex SQL statements. First we create a cursor, then open it using OPEN statement. After that we fetch data using FETCH command till it reaches end of result set. To close a cursor use CLOSE command. Schema is a name for a data structure, such as a table or view, that contains some piece of data. Within a database there might be many schemas, each with its own set of tables, views, indexes, and so forth. There are three kinds of schemas in SQL Server; User Schemas, System Schemas, and Default Schema. Database Blackbox testing means, testing in a way so as not to change the black box. The tester is unaware of internal implementation details, such as algorithms. He/she interacts with the application through a standardized interface, usually using standard input output format. Using this method, tester does not get confused by the complexity of the underlying code logic. Data Integrity means ensuring that a record that is changed is validated against other records such that errors cannot occur if a transaction conflicts with another transaction. For example, consider a company where sales staff enter customer orders into the system. If an order is entered incorrectly, it must still be possible to cancel it without causing damage to other customers’ information. A transactional update will ensure that this happens. RLS helps us to restrict user’s access to objects in a Database. It uses security level in place of roles to control access to various objects. We have 3 major categories of Access Rights as listed below: • Ownership – This right gives privilege to owner(user) to perform activities on the Object. • Membership – This right gives permission to users who can see rows owned by the user. • Expressions – This rights give scope over what expressions are allowed to execute in the context of the object. This right restricts how functions may be used. Normally, a normalization rule describes a relational model. Using entity-relationship diagrams, rules are formulated. These rules define all the relationships between entities. In general rule two things are kept in mind while designing a database. That is, firstly only one thing from different group must be related together, meaning that different groups should be stored within separate tables. Secondly every row within a given table should contain only unique value.Your database design is perfect if these two conditions are met. When you need to join more than one source table together to produce a single final output, instead of using subqueries. An example would be joining multiple tables containing employee salary and benefits. ERD is a tool to develop a conceptual view of a database. It allows people to graphically depict relations among database entities. A simple ERD shows how objects relate and why they have been organized that way. A relationship might represent any number of attributes, including foreign key columns, properties or even business rules. You may also have many-to-many relationships in which there are several objects linking each other. It replaces NULL values with some specified string in case when expression returns NULL. It is basically a replacement for IFNULL() statement. Instead of checking if the column contains null value we just substitute this value with some default value as defined in coalesce_default parameter of this function. SQL injection attacks occur when users provide input in such a way that enables them to manipulate the result set produced by the query. For e.g. suppose the user enters ‘;DROP TABLE Students’ in place of name field. This will cause drop table statement to execute against selected student database. Subquery is a type of SELECT statement. There are three types of subqueries • Correlated Subquery: Here the results of one or more queries are returned along with the main query. • Uncorrelated Subquery: Here no data is returned but only the query itself is executed. • Scalar Subquery: Here the entire query is evaluated and the resulting value is returned. Both of these statements insert same data into the target table. But the former inserts the data without specifying the column names whereas the latter specifies the column names. Group functions help us to perform aggregate functions like SUM(), AVG(), MAX(), MIN(). They can be used to find out the maximum/minimum value of a particular column in a table. A relationship is a connection between two or more tables. It helps us to link two tables together. For instance, if we want to know about students who have taken a course, we could create a new table called CoursesStudents where we store information about the courses taken by the students. Then we can link this table to the Student table using a foreign key. The following are the different types of relationships: • One To Many: When one object has many associations with another object. • Many To Many: When one or more objects have many associations with another object(s). • One To One: When one object has exactly one association with another object. • Many To One: When one or more object has exactly one association (or reference) with another object. Merge statement is used to merge multiple rows from source table into single row in destination table. It also works on the columns which are not part of primary key. With our courses, you’ll have the knowledge necessary to shine during an interview, from basic syntax to advanced topics. Plus, our interactive classes and practice sql interview questions and answers for freshers will help you drill down and hone your skills even further. So what are you waiting for? Enroll today at FITA Academy and start preparing for your next SQL interview! Recursive stored procedures are similar to procedural programming languages. They can be used to calculate complex operations. Suppose you have a requirement to calculate the sales tax for a particular product. You would write a stored procedure to do so. Now the problem arises that you don’t want to repeat the calculation every time your application needs to calculate the sales tax. Hence, you might think of creating a stored procedure which calculates the sales tax at run-time. Similarly, you can use recursive stored procedures to solve problems related to calculations of taxes, interest etc. Triggers are a mechanism which automatically executes certain actions when certain events happen. For e.g., whenever there is any update operation on a record, trigger will automatically recalculate the price of an item. There are three types of triggers: • AFTER TRIGGER – This type of trigger runs after the action specified in it completes successfully. • BEFORE TRIGGER – This trigger runs before the specified action takes place. • INSTEAD OF TRIGGER – This kind of trigger runs instead of the specified action. Clause is a set of instructions which define the meaning of expressions in a query. Clauses are enclosed within parentheses (). The most common clauses are SELECT, WHERE, FROM, HAVING, ORDER BY, LIMIT, UNION, JOIN. HAVING clause filters out only those records which satisfy all the conditions defined inside the HAVING clause. Whereas WHERE clause filters out only those values which satisfy the condition stated in the WHERE clause. If both clauses are present then they are combined logically. Group By clause groups data based on some criteria. For example, suppose we have a database containing details of employees working in a company. We may group these employees based on their department. By using an alias, we can indicate the ordinal position of a column in a select list. A commas separate the alias name from the AS keyword to create an alias. For eg: SELECT emp_name FROM employee • SELECT emp_name FROM person • SELECT p.emp_name AS emp_name FROM persons P Aggregate function returns a measure of central tendency or a summary statistic of a collection of numeric data. Examples of such functions include AVG(), MAX(), MIN() etc. The most commonly used aggregates are SUM(), COUNT(), AVG(). Scalar functions return one value. These functions are very useful as they can be used to perform various mathematical operations like addition, subtraction, multiplication, division etc. On the other hand, non-scalar functions return more than one value. For example, using a scalar function, SUM(), you can add all the numbers entered by users. A non-scalar function is also known as an aggregate function. Distinct command removes duplicate rows from a table. It operates on two tables joined with each other. A distinct operator works on columns of the same name. If you have a table named EMP having columns NAME and DEPARTMENT, then you can apply DISTINCT on this table to remove duplicate names. Null IF is used to check whether a given expression equals null. So, if you run this command without specifying any expression, it checks for null values. A view is a virtual table that provides read-only access to another table. Views provide fast access to data but cannot be updated directly. They are similar to queries except that views are stored in memory whereas queries are saved on disk. In order to modify results of a query, we first create a new query by modifying the original query. Then we save this new query to a file so that later we can retrieve the modified result set. Using a view, we don’t need to go through these steps. To create a view, we simply run a CREATE VIEW statement. Collations are rules that help determine how strings are compared within the same database. Different collations can cause strings to sort differently. Each character has a collating sequence associated with it. This sequence determines how the string should be sorted. There are many different types of collations available. When we talk about collation sensitivity, we mean the ability of some characters to dictate their relative positions in comparison with others. Some characters are more sensitive than others. That means that when comparing two strings, certain characters may influence the sorting priority. Characters that might affect the sorting criteria are termed as primary key characters. Local variables are temporary variable which exist only inside an execution context. We can declare local variables at any point within our script. Variables declared outside of an execution context are called global variables. It is the entire program or module that is covered by global variables. Throughout the application, they are accessible. An auto increment column automatically increments its value whenever a row is inserted into the table. The default behavior of auto incrementing is to start counting at 1. You can change the starting value by setting the AUTO_INCREMENT property. PL/SQL is a database programming language. It was invented by Oracle Corporation. In contrast, SQL is a standard protocol and language for querying databases. SQL is generally used for retrieving data from a relational database. Use the following query to get the total number of records in a particular table. SELECT COUNT() FROM TABLE_NAME; A datawarehouse (DW) is a collection of information that is organized in such a way that it can be accessed quickly using a variety of tools. DWs are usually designed to store large amounts of data, often terabytes or even petabytes. Create the tables with identical structures. For example, if you want to copy the structure of one table to another then use the following command. ALTER TABLE t1 RENAME TO t2; Copy data from one table to another table using INSERT INTO… SELECT syntax. For example, insert rows from one table to another using the following command. INSERT INTO t1 SELECT * FROM t2; Comments are special sequences of characters enclosed in /* */ that appear anywhere in a SQL statement. They do not have any effect on the operation of the statement. However, they are useful for documenting statements. Rank function returns the position of a given row in a group. It works on ordered sets of values. On the other hand, dense rank function gives the position of a row in relation to all the rows in the set. Row_number() is used to generate sequential numbers for each row in a result set. This is used to order result set in ascending or descending order. Rank() is used to assign a rank to each row in a result set based on the specified ordering. Ordering by columns without specifying direction is called ascending order. In descending order, the last column specified will be first. If no column is specified, the result is sorted alphabetically. We hope this article has helped you understand what SQL Interview Questions are, What are SQL Query Languages like SQL, MS-SQL, MySQL etc. The FITA Academy offers online training and certification for SQL. The academy’s curriculum is designed to help candidates increase their skills for both technical and non-technical SQL questions. The academy teaches how to construct effective queries, understand data types, use query optimization techniques, and respond to common SQL interview questions. FITA Academy also offers group training and individual coaching for those who need extra assistance. A well-prepared candidate will be able to answer questions on topics such as data types, querying techniques, and database design. The FITA Academy offers a comprehensive course that can help you prepare for a successful SQL interview. Supplementary Resources SQL is a powerful and widely used database management system that helps database administrators manage data. If you’re looking to get a job in the database field, be prepared to take some SQL interview. Here are some resources to help you study for those exams: The PHP and MySQL Interview Questions and Answers resource offers interview questions and answers for PHP and MySQL, respectively. It’s an excellent place to start if you’re new to SQL and want to improve your skills. It’s a standard for data manipulation and retrieval. Consequently, SQL is in high demand for many positions in companies. The popularity of SQL means that there are many resources available to help you prepare for your SQL interview. This includes studying Oracle Interview Questions and Answers to prepare for your Oracle interview. However, most Oracle interviews are about database administration and storage. So FITA Academy gives you the best SQL training, so that you can ace your interview and get placed. It is important that you check out the Freshers Salary page in order to increase your chances of landing a SQL job interview. You can find a wealth of information on all things related to salary, from what to expect in terms of pay rates to what to expect when it comes to benefits. Quick Enquiry Please wait while submission in progress... 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Charles Babbage Biography of Charles Babbage Charles Babbage, credited deservedly as Father of the Computer, the world renowned inventor of Differential Engine and Analytical Engine, was born on the 26th of December, 1791, in the family home at 44 Crosby Row, Walworth Road, London. He was the first child of Benjamin Babbage (1753-1827) and Elizabeth Plumleigh Babbage, née Teape (1759-1844). Benjamin Babbage Sr., Charles' grandfather, was mayor of Totnes in Devon (in southwest England). His son, also Benjamin (Charles' father) had started out as a goldsmith in the small town of Totnes, a picturesque port in Devonshire, and later became a successful merchant and banker, who had waited until he was 38 year of age and wealthy before marrying and moving to London in 1790 to join a new banking firm—Bitton Estate in Teignmouth. Elizabeth Teape came from a prominent Devonshire family. After Charles, the family had two other sons, who died in infancy, and a daughter—Mary Ann (born 1798). She outlived Charles and the two siblings remained close throughout their lives. Young Charles was brought up as an Anglican and received his earliest education at home. His childhood was marred by chronic illness and hoping that country living would improve his health, around the age of eight his parents began to sent him to country schools. In 1803, his family returned to Devon, and in improved health, Charles was sent to a small residential school in the village of Enfield near London, where he remained for three years. The teacher at Enfield was Stephen Freeman, an amateur astronomer and namely he awakened Charles’s interest in science and mathematics. Charles then moved to a small school near Cambridge for a couple of years. This may have been to prepare for entrance to the University of Cambridge, but it made little impression on him. At age 16 or 17, Babbage returned to Devon to live with his parents. He learned Latin and Greek with a tutor and also spent much time studying mathematics on his own. By then, he was passionately fond of algebra and devoured every book he could find on the subject. In 1808, the Babbage family moved into the old Rowdens house in East Teignmouth, and Benjamin Babbage became a warden of the nearby St. Michael’s Church. In October 1810, Charles Babbage began his studies at Trinity College, Cambridge. It meant new perspectives and he found the environment, the books and social life intensely stimulating. Here he was to meet new friends who would remain close to him for the rest of his life. His days were spent in sampling the pleasures of undergraduate existence parties with plenty of good food and drink, Sunday breakfasts with his friends after Church, chess and games of whist and trips on the Cam. There was a servant to take care of the routine chores and make Babbage's life all the more agreeable. All this was financed with the 300 pounds, a big sum for the time, which Charles received as an annual allowance from his father. Among his new friends, John Frederick William Herschel soon took first place. (He was the son of William Herschel, the outstanding astronomer, who had discovered the planet Uranus in 1781. John followed in his father’s footsteps, and became one of the leading men of science in England during the 1800s.) Together they began to devote themselves to mathematics. Other friends of Babbage were the famous mathematician George Peacock and Edward Ryan, a famous English lawyer. In 1812, Babbage, Peacock, Herschel and some other students had founded a little association called the Analytical Society. Its purpose was to introduce continental mathematical methods into the conservative Cambridge. Georgiana BabbageIn the spring of 1814, Charles Babbage received his B.A. Shortly afterwards, on 25th July 1814, he married to Georgiana Whitmore (1792-1827) (see the nearby portrait), one of the 8 daughters of a wealthy Shropshire family and in the fall, they moved to London. The marriage was not welcomed by his father Benjamin and it would appear that the relations between father and son were far from harmonious. Benjamin Babbage had no complaints against Georgiana. His attitude was that, like himself, Charles should wait until he was properly established financially. Fortunately, Benjamin continued his £300 annual allowance, to which Georgiana could add £150 of her own. With such an income, the couple could maintain a modest life without lavish entertaining. In the August next year was born his first son—Benjamin Herschel (1815-1878). Charles and Georgiana had eight children, 7 sons and 1 daughter, but only three—the above-mentioned Benjamin Herschel, 5th son, Dugald Bromheald (1823-1901), and 6th son, Henry Prevost (1824-1918) survived to adulthood. All other children died young. In 1815 Babbage becomes a member of the Royal Society. For a while, he sought paid employment, to prove to his father that he could make something of himself. In 1816, he applied for the post of math professor at East India College at Hartford, but his hopes were soon dashed when another candidate was chosen instead. In 1819, Babbage once more applied for a professorship, this time at Edinburgh. But despite all his recommendations from prominent French and English mathematicians, Babbage did not gain the position. At the same time, he also applied for a seat on the Board of Longitude but this too ended in failure. In 1820, he made a new application but to no avail. In 1819 Babbage travels to Paris to visit French scientists. There he gets first inspiration for Difference Engine from Baron Gaspard de Prony’s use of division of labor for calculating tables. Charles Babbage as youngAt the close of 1820, Charles Babbage by now twenty-nine was still without any profession. For the previous six years, he had tried to find something suitable. He had carried out intensive mathematical research and had published a fair number of articles. He had presented several of his findings in lectures at the Royal Society, among whose illustrious members he had managed to establish himself. He had also, once again, shown his predilection for reform by becoming one of the cofounders of the Astronomical Society in 1820. Despite of his unsuccessful attempts to find a job, the family seemed to manage quite comfortably financially. In 1814 Babbage made his first step in the field of engineering. He invented a new type of lock which he was interested in having manufactured. This was possibly his first serious excursion into that area of human endeavor, which was with time to cast its spell on him. The real passion about engineering will possess him in 1821, when he will begin his Difference Engine. Besides his lifetime engagement with the construction of Differential Engine and Analytical Engine, Babbage did make occasional forays into other fields. In 1824, he was invited by some investors to organize a life insurance company. The new challenge intrigued him, and he threw himself into the task of determining the appropriate rates to charge for life insurance policies. Having collected a lot of information, Babbage decided that he would have to make some other use of it. In 1826, he published a book on the life insurance industry, A Comparative View of the Various Institutions for the Assurance of Lives. In fewer than 200 pages, this book provided a very useful consumer’s guide to the life insurance companies in England at that time. Readers could use it to compare companies and make intelligent decisions about which one would suit their particular needs. In 1827 Babbage decided to publish tables of logarithms. He compared several tables, published since then in England. Wherever they differed, he recalculated the value so that he could produce a table completely free from error. With the help of an army engineer, he directed the work of a number of clerks. The corrected table was published in 1827. This table was reprinted many times, even after 1900. Babbage's Table of Logarithms of the Natural Numbers from 1 to 108,000 was a paradigm of accuracy and was extensively used into the twentieth century. The year 1827 was a devastating year for Charles. In February of 1827, his father died in Devon at the age of 73. Old Benjamin left sufficient funds to care for his wife, Betty, who moved to London to live with Charles and his family. Babbage inherited about £100,000, the bulk of his father's estate, which made him a very rich man. The interest on the investments and the rent on the properties provided a comfortable income for the rest of his life. However, his view of a comfortable life did not last long. In July of the same year, his son Charles Whitmore was struck with a childhood disease and died at the age of 10. Then, in a month later, Charles’s wife Georgiana contracted a serious illness. On the 1st of September, both she and a newborn son Alexander Forbes also died. These deaths caused Babbage to go into a mental breakdown which delayed the construction of his machines. In 1828, Charles was elected as Lucasian Professor of Mathematics at Cambridge. This university chair, once held by Isaac Newton, was a great honor, though it carried an annual salary of less than £100. Babbage however did not think it was worth the distraction from his beloved Difference Engine. He held the post for ten years, however, he did not live in Cambridge and seldom lectured there. Nevertheless, he was always grateful for the appointment, which he called the only honour I ever received in my own country. At the end of 1827 Babbage made a long (one year) trip to Europe, when he met a lot of leading European scientists. After returning to England, from 1829 to 1834, Babbage engaged in electoral politics, promoting candidates and even standing for election himself. In 1830 he published a book—Reflections on the Decline of Science in England, which is the best known of Babbage’s many polemics against the scientific institutions of his day and fueled much debate at the time and after. In addition to the affairs of his family, he continued with the Difference Engine, and managed to write a book on the economy of manufacturing. He began to also to hold regular Saturday evening parties, initially in order to introduce his teenage children, Herschel and Georgiana, into society. Before long, the Babbage soirées formed an important part of the London social scene. Often, the guest list exceeded 200. They came from all parts of polite society: lawyers and judges, doctors and surgeons, deacons and bishops, and scholars and artists by the score. In the midst of this full bustle of activity during the 1830s, personal tragedy again struck Babbage. In 1834, his beloved daughter Georgiana became ill and died. She was only 17 years old. To deal with his grief, Charles threw himself more deeply into his work. Babbage was able to turn every experience to advantage. After all his visits to workshops and factories both in England and on the continent, he sought to draw general principles from them. In 1832, Babbage compiled these principles into the more than 30 chapters of his book On the Economy of Machinery and Manufactures. Within three years, there were four editions in England, one in America, and translations into German, French, Italian, Spanish, Swedish, and Russian—a real best-seller. Charles Babbage at middle ageDuring the 1830s Babbage became progressively more involved in developing the efficiency of rail transport in England. Charles and his son Herschel even constructed a special device, something like black box, which measured the speed of the train and its degree of vibration. In 1842 the oldest son of Babbage—Herschel, with his family and his brother Dugald, went off on a railway project in Italy. After other jobs, these two sons went to Australia in 1851 to conduct a geological survey. The third son, Henry, decided to join the Indian army. He took up his post there in 1843. Charles’s mother, Betty, was left alone in the old house, where she died in 1844 in her mid-eighties. Charles fell into a routine that lasted most of the rest of his life. He devoted mornings and afternoons to writing or work on the Analytical Engine or Differential Engine, and then evenings to dinner, followed by a party, a play, or the opera. In 1861, at the age of 70, Charles Babbage became more aware of his own mortality. He began to devote part of his time to writing a collection of reminiscences. His autobiography, titled Passages from the Life of a Philosopher, was published in 1864. Babbage was a quite eccentric and a man of extremes. His friends could do no wrong and his enemies could do no right. He once contacted the poet Alfred Tennyson in response to his poem "The Vision of Sin". Babbage wrote, "In your otherwise beautiful poem, one verse reads, Every moment dies a man, Every moment one is born. ...If this were true, the population of the world would be at a standstill. In truth, the rate of birth is slightly in excess of that of death. I would suggest that the next version of your poem should read: Every moment dies a man, Every moment 1 1/16 is born. Strictly speaking, the actual figure is so long I cannot get it into a line, but I believe the figure 1 1/16 will be sufficiently accurate for poetry." Babbage especially hated street music, and once wrote: It is difficult to estimate the misery inflicted upon thousands of persons, and the absolute pecuniary penalty imposed upon multitudes of intellectual workers by the loss of their time, destroyed by organ-grinders and other similar nuisances. It is said that organ grinders were playing deliberately outside his house on the day he died. Charles Babbage oldThere is no doubt, that Babbage was an outstanding genius and he is the only man (if any!), who deserves the title Father of the Computer. His Differential Engine was a sophisticated specialized calculating machine, try to compare this monster with the ubiquitous calculator of Colmar, manufactured at the same time. But Differential Engine was nothing, compared to the Analytical Engine. It's unbelievable, that Babbage dared to design a universal computer more than 100 years, before such a machine to be produced. And he was capable to make it, if he had the necessary support. 100 years in the field of computers is equal to 1000 years in many other areas. At the same time his striving for perfection and his difficult personality foiled a lot of his great ideas. After 10 years of work and huge sum of money spent, he was very close to completion of Differential Engine, but his disputes with the engineer Clement, and some other trammels, spoiled his plans. Despite his many achievements, the failure to construct his calculating machines left Babbage in his declining years a disappointed and embittered man. He died of renal inadequacy, secondary to cystitis at age 79 on 18th October 1871 at his home on Dorset Street, London, and was buried in London's Kensal Green Cemetery. Babbage's brain is preserved at the Science Museum in London. Charles Babbage in 1848 Charles Babbage (a painting by Samuel Laurence from 1845, at the National Portrait Gallery in London) Charles Babbage was a genius of the first order—a mathematician and professor, an engineer and inventor, a politician, a writer, a cryptographer, a founder of scientific organizations, and an expert on industry. His pioneering book, On the Economy of Machinery and Manufactures, was cited repeatedly by Marx in Capital and by John Stuart Mill in Principles of Political Economy. He was a human dynamo who needed only five or six hours of sleep a day and who was driven by a millennarian vision of man and machine that brought him within a hair's breadth of the invention of the greatest of machines—the computer. There are only two ways to live your life. One is as though nothing is a miracle. The other is as though everything is a miracle. Albert Einstein
A public repository on Docker Hub offers the latest versions of the Satellite. If you want to use a Helm chart to install Satellites to docker, follow these instructions. You configure the Satellites using environment variables. Please see Satellite Configuration Parameters for a full listing and expanded explanations of variables and recommended settings. You can also use the following template and fill in your own values for the variables (defaults are entered for you, but you can change them). If you want to use yaml instead of the command line, you can use the yaml file in the AWS configuration topic. If you are happy with the default port numbers and don’t need TLS, you only need to modify the following settings: * satellite_key * pool * bytes_per_project 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 docker run -e COLLECTOR_SATELLITE_KEY= -e COLLECTOR_POOL= -e COLLECTOR_GUID= -e COLLECTOR_STATSD_HOST= -e COLLECTOR_STATSD_PORT= -e COLLECTOR_STATSD_EXPORT_STATSD= #-----Only if using Datadog for Satellite Metrics---- -e COLLECTOR_STATSD_EXPORT_DOGSTATSD= -e COLLECTOR_STATSD_DOGSTATSD_TAGS= #----------------------------------------------------- -e COLLECTOR_STATSD_PREFIX=lightstep -e COLLECTOR_STATSD_SATELLITE_PREFIX=satellite -e COLLECTOR_STATSD_CLIENT_PREFIX=client -e COLLECTOR_DIAGNOSTIC_PORT=8000 -p 8000:8000 -e COLLECTOR_ADMIN_PLAIN_PORT=8180 -p 8180:8180 -e COLLECTOR_ADMIN_SECURE_PORT= 8181 -p 8181:8181 -e COLLECTOR_HTTP_PLAIN_PORT=8182 -p 8182:8182 -e COLLECTOR_HTTP_SECURE_PORT=8183 -p 8183:8183 -e COLLECTOR_GRPC_PLAIN_PORT=8184 -p 8184:8184 -e COLLECTOR_GRPC_SECURE_PORT=8185 -p 8185:8185 -e COLLECTOR_PLAIN_PORT=8186 -p 8186:8186 -e COLLECTOR_SECURE_PORT=8187 -p 8187:8187 -e COLLECTOR_REPORTER_BYTES_PER_PROJECT=1000000000 -e COLLECTOR_REPORTER_BYTES_PER_PROJECT_OVERRIDES= ?? -e COLLECTOR_TLS_CERT_PREFIX= ?? -e COLLECTOR_DISABLE_ACCESS_TOKEN_CHECKING= -e COLLECTOR_PROJECT_NAME= -e COLLECTOR_INGESTION_TAGS= lightstep/collector:latest Here is an example using a Docker Compose file. This configuration sends Spans over http on port 8182 with secure traffic configured to go over port 8183. Change the COLLECTOR_API_KEY to your satellite key for your project, save it as docker-compose.yml and then do docker-compose up. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 version: '3' services: satellite: image: lightstep/collector:latest environment: - COLLECTOR_SATELLITE_KEY=someLongRandomStringOfCharacaters - COLLECTOR_DIAGNOSTIC_PORT=8000 - COLLECTOR_ADMIN_PLAIN_PORT=8180 - COLLECTOR_HTTP_PLAIN_PORT=8182 - COLLECTOR_HTTP_SECURE_PORT=8183 - COLLECTOR_REPORTER_BYTES_PER_PROJECT=1000000000 ports: - "8000:8000" #Diagnostics - "8180:8180" #COLLECTOR_ADMIN_PLAIN_PORT, Required for health checks - "8182:8182" #COLLECTOR_HTTP_PLAIN_PORT, Unsecure traffic, or secure traffic that terminates it's secure status before it hits the satellite - "8183:8183" #COLLECTOR_HTTP_SECURE_PORT, for Secure traffic After startup, check the diagnostics page (http://\{satellite\_host\}:8000/diagnostics) to view the Satellite configuration. When starting from command line as in the above example, the configuration will also be directed to stdout, which you can inspect for troubleshooting if the diagnostics page isn’t reachable. Follow these instructions to further validate your installation and troubleshoot any issues. You configure your Lightstep tracers to communicate with your Satellites by providing their location. If you are using a single Satellite, you provide the DNS name or IP address of that Satellite. For more than one, use the name or address of the load balancer you’ve deployed. Find out how to do that in the language-specific Quick Start guides. By default, client libraries use secure connections. If you’ve configured your Satellites to use plainport connections, you must configure your tracer appropriately.
Need advice about which tool to choose?Ask the StackShare community! Apache Beam 164 315 + 1 14 Apache Flink 443 720 + 1 36 Add tool Advice on Apache Beam and Apache Flink Nilesh Akhade Technical Architect at Self Employed · \| 5 upvotes · 301.8K views We have a Kafka topic having events of type A and type B. We need to perform an inner join on both type of events using some common field (primary-key). The joined events to be inserted in Elasticsearch. In usual cases, type A and type B events (with same key) observed to be close upto 15 minutes. But in some cases they may be far from each other, lets say 6 hours. Sometimes event of either of the types never come. In all cases, we should be able to find joined events instantly after they are joined and not-joined events within 15 minutes. See more Replies (2) Recommends ElasticsearchElasticsearch The first solution that came to me is to use upsert to update ElasticSearch: 1. Use the primary-key as ES document id 2. Upsert the records to ES as soon as you receive them. As you are using upsert, the 2nd record of the same primary-key will not overwrite the 1st one, but will be merged with it. Cons: The load on ES will be higher, due to upsert. To use Flink: 1. Create a KeyedDataStream by the primary-key 2. In the ProcessFunction, save the first record in a State. At the same time, create a Timer for 15 minutes in the future 3. When the 2nd record comes, read the 1st record from the State, merge those two, and send out the result, and clear the State and the Timer if it has not fired 4. When the Timer fires, read the 1st record from the State and send out as the output record. 5. Have a 2nd Timer of 6 hours (or more) if you are not using Windowing to clean up the State Pro: if you have already having Flink ingesting this stream. Otherwise, I would just go with the 1st solution. See more Akshaya Rawat Senior Specialist Platform at Publicis Sapient · \| 3 upvotes · 192.3K views Recommends Apache SparkApache Spark Please refer "Structured Streaming" feature of Spark. Refer "Stream - Stream Join" at https://spark.apache.org/docs/latest/structured-streaming-programming-guide.html#stream-stream-joins . In short you need to specify "Define watermark delays on both inputs" and "Define a constraint on time across the two inputs" See more Get Advice from developers at your company using Private StackShare. Sign up for Private StackShare. Learn More Pros of Apache Beam Pros of Apache Flink • 5 Open-source • 5 Cross-platform • 2 Portable • 2 Unified batch and stream processing • 15 Unified batch and stream processing • 8 Easy to use streaming apis • 8 Out-of-the box connector to kinesis,s3,hdfs • 3 Open Source • 2 Low latency Sign up to add or upvote prosMake informed product decisions - No public GitHub repository available - What is Apache Beam? It implements batch and streaming data processing jobs that run on any execution engine. It executes pipelines on multiple execution environments. What is Apache Flink? Apache Flink is an open source system for fast and versatile data analytics in clusters. Flink supports batch and streaming analytics, in one system. Analytical programs can be written in concise and elegant APIs in Java and Scala. Need advice about which tool to choose?Ask the StackShare community! What companies use Apache Beam? What companies use Apache Flink? See which teams inside your own company are using Apache Beam or Apache Flink. Sign up for Private StackShareLearn More Sign up to get full access to all the companiesMake informed product decisions What tools integrate with Apache Beam? What tools integrate with Apache Flink? Sign up to get full access to all the tool integrationsMake informed product decisions Blog Posts What are some alternatives to Apache Beam and Apache Flink? Apache Spark Spark is a fast and general processing engine compatible with Hadoop data. It can run in Hadoop clusters through YARN or Spark's standalone mode, and it can process data in HDFS, HBase, Cassandra, Hive, and any Hadoop InputFormat. It is designed to perform both batch processing (similar to MapReduce) and new workloads like streaming, interactive queries, and machine learning. Kafka Streams It is a client library for building applications and microservices, where the input and output data are stored in Kafka clusters. It combines the simplicity of writing and deploying standard Java and Scala applications on the client side with the benefits of Kafka's server-side cluster technology. Kafka Kafka is a distributed, partitioned, replicated commit log service. It provides the functionality of a messaging system, but with a unique design. Airflow Use Airflow to author workflows as directed acyclic graphs (DAGs) of tasks. The Airflow scheduler executes your tasks on an array of workers while following the specified dependencies. Rich command lines utilities makes performing complex surgeries on DAGs a snap. The rich user interface makes it easy to visualize pipelines running in production, monitor progress and troubleshoot issues when needed. Google Cloud Dataflow Google Cloud Dataflow is a unified programming model and a managed service for developing and executing a wide range of data processing patterns including ETL, batch computation, and continuous computation. Cloud Dataflow frees you from operational tasks like resource management and performance optimization. See all alternatives
Tell me more × Stack Overflow is a question and answer site for professional and enthusiast programmers. It's 100% free, no registration required. I want to create a UserControl in WPF through which I want to expose a collection property. I want to change the UI of the UserControl based on the changes in collection. For example, lets say I have a collection of strings which is binded to my UserControl. Based on that collection i want to create buttons on the UserControl containing those text as button text. Is there a way I can acieve this? share\|improve this question add comment 1 Answer up vote 1 down vote accepted Yes, you can set a DataTemplate containing a button for an ItemsControl control that is binded to that collection. For Example: //For code: items.DataContext = new List<string> { "Item 1", "Item 2", "Item 3" }; //For XAML <ItemsControl x:Name="items" ItemsSource="{Binding}"> <ItemsControl.ItemTemplate> <DataTemplate> <Button Content="{Binding}" /> </DataTemplate> </ItemsControl.ItemTemplate> </ItemsControl> share\|improve this answer Thanks very much. – deepak May 25 '09 at 8:25 add comment Your Answer discard By posting your answer, you agree to the privacy policy and terms of service. Not the answer you're looking for? Browse other questions tagged or ask your own question.
Using iMessage on iPhone and iPad Discussion in 'iOS 5 and earlier' started by OneForGadgets, Oct 16, 2011. 1. OneForGadgets macrumors member Joined: Sep 1, 2010 #1 Is anyone else having problems with transferring from their iPhone to iPad with iMessage? I will be in the middle of a conversation and move to my iPad from my iPhone and a new thread will start with the person I am talking to. I know on the iPhone it uses your phone number as the default and your email address for iPad, but I wold like to just use the email address instead. Anyone else having this problem? 2. Transeau macrumors 6502a Transeau Joined: Jan 18, 2005 Location: Alta Loma, CA #2 You need to insure that the Caller ID on both the iPad and iPhone is set to your Apple ID. Most issues are with the phone being set to the phone number and not the Apple ID 3. GeorgiaApple macrumors newbie GeorgiaApple Joined: Jul 5, 2011 Location: Georgia, Tbilisi #3 Hmm. I think iMassage doesnot work with CDMA. because it requires Wifi or 3G ... 4. RolledUp20s macrumors regular Joined: Jun 14, 2011 Location: Wrexham, North Wales #4 Lol, i found it annoying that having a convo on my ipad would send everything to my iphone aswel, so ive set up a seperate email for my ipad now. But to get that to work like you want i had ipad n iphone imessages to set to recieve at the same email. ? Think that was it.. 5. miamialley macrumors 68030 miamialley Joined: Jul 28, 2008 Location: Los Angeles, CA #5 There are a few threads about this. I've got the caller ID set up and it still does not always work properly. And it has nothing to do with CDMA. It seems buggy playing between devices for the same user. http://forums.macrumors.com/showthread.php?t=1251006&highlight= 6. OneForGadgets thread starter macrumors member Joined: Sep 1, 2010 #6 Wow, I didn't think that many people were having problems. Hopefully they have a work around because asking for someone's Apple ID to prevent this is just stupid. Share This Page
fredrik fredrik - 11 months ago 63 Python Question How to make chocolatey install python2 into custom path? I'm installing Python 2.7.11 using Chocolatey, which installs into C:\tools\python2 : choco install python2 -y Is there any way I can get Python to install into C:\Python27 instead? Answer Yes, override the install arguments - choco install python2 -y -o -ia "'/qn /norestart ALLUSERS=1 TARGETDIR=c:\Python27'" Alternatively you can create a Python27 folder and it will install there. Note that is determined by reading through the chocolateyInstall.ps1 script - https://chocolatey.org/packages/python2#files
How To Make Use of Unstructured Data in SOC Detection Workflows Security operations have become increasingly proficient at using structured threat intelligence to enrich alerts, and accelerate investigation and threat hunting workflows. Threat intelligence platforms have largely automated the process of collecting, extracting and normalizing intel from structured data sources. But a significant amount of intelligence available today is still shared through blogs, advisories and research articles, which requires tedious manual processes to make it usable by SIEM and SOAR tools. Researchers and practitioners have been working on challenges related to extracting unstructured intelligence and making it useful for a variety of use cases. For those interested in this topic and are looking for a starting point I have created a brief list of blogs, projects and presentations covering different approaches and related NLP methods. This is not meant to be a long list of articles - it is meant to be a starting point that can help you drill down further. Frameworks like STIX and Mitre ATT&CK offer an intermediary translation step between unstructured intelligence and machine usable intelligence. Advances in NLP and deep learning techniques will also spur new ideas and approaches to solving this problem. As new work comes to light I will keep adding to this list - and always happy to get recommendations from you!
OpenBSD manual page server Manual Page Search Parameters SEMOP(2) System Calls Manual SEMOP(2) semopsemaphore operations #include <sys/sem.h> int semop(int semid, struct sembuf sops, size_t nsops); () provides a number of atomic operations on a set of semaphores. The semaphore set is specified by semid. sops is an array of semaphore operations, nsops is the number of operations in this array. The sembuf structures in the array contain the following members: u_short sem_num; / semaphore # / short sem_op; / semaphore operation / short sem_flg; / operation flags */ Each operation (specified in sem_op) is applied to semaphore number sem_num in the set of semaphores specified by semid. The value of sem_op determines the action taken in the following way: The behavior of each operation is influenced by the flags set in sem_flg in the following way: In the case where the calling process would normally block, waiting for a semaphore to reach a certain value, IPC_NOWAIT makes the call return immediately, returning a value of -1 and setting errno to EAGAIN. Keep track of the changes that this call makes to the value of a semaphore, so that they can be undone when the calling process terminates. This is useful to prevent other processes waiting on a semaphore to block forever, should the process that has the semaphore locked terminate in a critical section. Upon successful completion, the value 0 is returned; otherwise the value -1 is returned and the global variable errno is set to indicate the error. semop() will fail if: [] There is no semaphore associated with semid. [] The semaphore set was removed while the process was waiting for one of its semaphores to reach a certain value. [] The calling process has no permission to access the specified semaphore set. [] The value of nsops is too big. The maximum is specified in MAX_SOPS in <sys/sem.h>. [] sem_num in one of the sem_buf structures is less than 0, or greater than the actual number of semaphores in the set specified by semid. [] was requested, and there is not enough space left in the kernel to store the undo information. [] The requested operation cannot immediately be performed, and IPC_NOWAIT was set in sem_flg. [] sops points to an illegal address. semctl(2), semget(2) November 1, 2015 OpenBSD-6.0
TitlePolyunsaturated fatty acids inhibit S14 gene transcription in rat liver and cultured hepatocytes. Publication TypeJournal Article Year of Publication1993 AuthorsJump, DB, Clarke, SD, MacDougald, O, Thelen, A JournalProc Natl Acad Sci U S A Volume90 Issue18 Pagination8454-8 Date Published1993 Sep 15 ISSN0027-8424 KeywordsAdipose Tissue, Animals, Base Sequence, Cells, Cultured, Chloramphenicol O-Acetyltransferase, Epididymis, Fish Oils, Gene Expression Regulation, Kinetics, Liver, Male, Molecular Sequence Data, Nuclear Proteins, Oligodeoxyribonucleotides, Oligonucleotides, Antisense, Polymerase Chain Reaction, Promoter Regions, Genetic, Protein Biosynthesis, Proteins, Rats, Rats, Sprague-Dawley, Recombinant Proteins, RNA, Messenger, Sequence Deletion, Transcription Factors, Transcription, Genetic, Transfection, Triolein Abstract Polyunsaturated fatty acids (PUFAs) have been shown to have significant effects on hepatic lipogenic gene expression. The S14 gene has been used as a model to examine the effects of PUFAs on hepatic lipogenic gene expression. In vivo studies showed that feeding rats a high carbohydrate diet containing menhaden oil rapidly (within hours) and significantly (> or = 50%) attenuates hepatic S14 gene transcription and S14 mRNA abundance. The suppressive effect of menhaden oil was both gene and tissue specific. The effect of PUFAs on expression of the S14 mRNA and a transfected S14 fusion gene (i.e., S14CAT4.3) was examined in cultured hepatocytes in the presence of triiodothyronine (T3), insulin, dexamethasone, and albumin under serum-free conditions. Whereas T3 stimulated both S14 mRNA (> 40-fold) and S14CAT4.3 (> 100-fold), eicosapentaenoic acid (C20:5 omega 3) significantly attenuated (> or = 80%) both S14 mRNA and S14CAT activity in a dose-dependent fashion. The effects of C20:5 on hepatocyte gene expression were both gene and fatty acid specific. Deletion analysis of transfected S14CAT fusion genes indicated that the S14 thyroid hormone response element (at -2.5 to -2.9 kb) was not sensitive to C20:5 control. The cis-linked PUFA response elements were localized to a region within the S14 proximal promoter (at -80 to -220 bp). This region also contains cis-acting elements that potentiate T3 activation of S14 gene transcription. These studies suggest that C20:5 (or its metabolites) regulates factors within the S14 proximal promoter region that are important for T3 activation of S14 gene transcription. DOI10.1073/pnas.90.18.8454 Alternate JournalProc. Natl. Acad. Sci. U.S.A. PubMed ID8378318 PubMed Central IDPMC47375 Grant ListR01 DK043220 / DK / NIDDK NIH HHS / United States DK43320 / DK / NIDDK NIH HHS / United States
Assessment \| Biopsychology \| Comparative \| Cognitive \| Developmental \| Language \| Individual differences \| Personality \| Philosophy \| Social \| Methods \| Statistics \| Clinical \| Educational \| Industrial \| Professional items \| World psychology \| Biological: Behavioural genetics · Evolutionary psychology · Neuroanatomy · Neurochemistry · Neuroendocrinology · Neuroscience · Psychoneuroimmunology · Physiological Psychology · Psychopharmacology (Index, Outline) microtubule-associated protein tau Symbol(s): MAPT DDPAC, MAPTL Locus: 17 q21 EC number [1] EntrezGene 4137 OMIM 157140 RefSeq NM_005910 UniProt P10636 Tau proteins are microtubule-associated proteins that are abundant in neurons in the central nervous system and are less common elsewhere. They were discovered in 1975 in Marc Kirschner's laboratory at Princeton University [Weingarten et al., 1975]. Tau proteins interact with tubulin to stabilize microtubules and promote tubulin assembly into microtubules. Tau has two ways of controlling microtubule stability: isoforms and phosphorylation. Six tau isoforms exist in brain tissue, and they are distinguished by their number of binding domains. Three isoforms have three binding domains and the other three have four binding domains. The binding domains are located in the carboxy-terminus of the protein and are positively-charged (allowing it to bind to the negatively-charged microtubule). The isoforms with four binding domains are better at stabilizing microtubules than those with three binding domains. The isoforms are a result of alternative splicing in exons 2,3, and 10 of the tau gene. Phosphorylation of tau is regulated by a host of kinases. For example, PKN, a serine/threonine kinase. When PKN is activated, it phosphorylates tau, resulting in disruption of microtubule organization [Taniguchi et al., 2001]. Hyperphosphorylation of the tau protein (tau inclusions), however, can result in the self-assembly of tangles of paired helical filaments and straight filaments, which are involved in the pathogenesis of Alzheimer's disease and other tauopathies [Alonso et al., 2001]. Tau protein is a highly soluble microtubule-associated protein (MAP). In humans, these proteins are mostly found in neurons compared to non-neuronal cells. One of tau's main functions is to modulate the stability of axonal microtubules. Tau is not present in dendrites and is active primarily in the distal portions of axons where it provides microtubule stabilization but also flexibility as needed. This contrasts with STOP proteins in the proximal portions of axons which essentially lock down the microtubules and MAP2 that stabilizes microtubules in dendrites. The tau gene locates on chromosome 17q21, containing 16 exons. The major tau protein in the human brain is encoded by 11 exons. Exon 2, 3 and 10 are alternative spliced, allowing six combinations (2-3-10-; 2+3-10-; 2+3+10-; 2-3-10+; 2+3-10+; 2+3+10+). Thus, in the human brain, the tau proteins constitute a family of six isoforms with the range from 352-441 amino acids. They differ in either no, one or two inserts of 29 amino acids at the N-terminal part (exon 2 and 3), and three or four repeat-regions at the C-terminal part exon 10 missing. So, the longest isoform in the CNS has four repeats (R1, R2, R3 and R4) and two inserts (441 amino acids total), while the shortest isoform has three repeats (R1, R3 and R4) and no insert (352 amino acids total). All of the six tau isoforms are present in an often hyperphosphorylated state in paired helical filaments from Alzheimer's Disease brain. In other neurodegenerative diseases, the deposition of aggregates enriched in certain tau isoforms has been reported. When misfolded this otherwise very soluble protein can form extremely insoluble aggregates that contribute to a number of neurodegerative diseases. References Alonso, A. del C., Zaidi, T., Novak, M., Grundke-Iqbal, I., Iqbal, K. (2001) Hyperphosphorylation induces self-assembly of tau into tangles of paired helical filaments/straight filaments. PNAS. (98) 6923-8. http://www.pnas.org/cgi/content/full/98/12/6923 Delacourte, A. (2005) Tauopathies: recent insights into old diseases. Folia Neuropathol (43) 244-257. http://www.new.termedia.pl/magazine.php?magazine_id=20&article_id=5368&magazine_subpage=FULL_TEXT Hirokawa, N., Shiomura, Y., Okabe, S. (1988) Tau proteins: the molecular structure and mode of binding on microtubules. J Cell Biol. (107) 1449-59. http://www.jcb.org/cgi/reprint/107/4/1449 Taniguchi, T., Kawamata, T., Mukai, H., Hasegawa, H., Isagawa, T., Yasuda, M., Hashimoto, T., Terashima, A., Nakai, M., Mori, H., Ono, Y., Tanaka, C. (2001) Phosphorylation of tau is regulated by PKN. J Biol Chem. (276) 10025-31. http://www.jbc.org/cgi/content/full/276/13/10025 Weingarten, MD., Lockwood, AH., Hwo, SY., Kirschner, MW. (1975) A protein factor essential for microtubule assembly. PNAS. (72) 1858-1862. http://www.pnas.org/cgi/content/abstract/72/5/1858 External links[edit \| edit source] de:Tau-Protein pl:Białko tau This page uses Creative Commons Licensed content from Wikipedia (view authors). Community content is available under CC-BY-SA unless otherwise noted.
Função gama Origem: Wikipédia, a enciclopédia livre. Ir para: navegação, pesquisa NoFonti.svg Este artigo ou se(c)ção cita fontes fiáveis e independentes, mas elas não cobrem todo o texto (desde Maio de 2012). Por favor, melhore este artigo providenciando mais fontes fiáveis e independentes, inserindo-as em notas de rodapé ou no corpo do texto, nos locais indicados. Encontre fontes: Googlenotícias, livros, acadêmicoScirusBing. Veja como referenciar e citar as fontes. A função gama nos números reais. Em matemática, a função gama é uma extensão da função factorial aos números complexos (após uma reparametrização).1 Esta função é definida por:1 \Gamma(z) = \int_0^\infty t^{z-1} e^{-t}\,\mathrm{d}t \,\! e verifica (para n natural): 1 \Gamma(n+1)=n!\,\! De modo geral: \Gamma(z+1)=z \ \Gamma(z)\, em particular: \Gamma\left(\frac{1}{2}\right) = \sqrt \pi\, Outra fórmula é a fórmula de reflexão de Euler, que diz que: \Gamma(1-z) \; \Gamma(z) = {\pi \over \sin{(\pi z)}} \,\! Prova [editar] É fácil perceber, através da regra da cadeia e de recursos da integração imprópria, que \int_0^\infty e^{-r}\,dr=\left.-e^{-r}\right\vert _0^\infty=1 Usando o método da substituição, de modo que r=st, dr=tds, (t>0 e fixo), obtém-se: \int_0^\infty e^{-st}\,ds=\frac{1}{t}, t>0 Derivando-se em relação a t e aplicando a fórmula de Leibniz: \int_0^\infty se^{-st}\,ds=\frac{1}{t^2}, t>0 Utilizando o mesmo processo novamente: \int_0^\infty s^2e^{-st}\,ds=\frac{1.2}{t^3}, t>0 Derivando sucessivas vezes em relação a t: \int_0^\infty s^{n}e^{-st}\,ds=\frac{n!}{t^{n+1}}, t>0 Para t=1 \int_0^\infty s^{n}e^{-s}\,ds=n! Dessa forma, tem-se uma função fatorial definida para quaisquer valores reais positivos x, de modo que: g(x)=\int_0^\infty e^{-s}s^{x}\,ds=x! Contudo, consagrou-se o uso de uma definição levemente destoante, a Função Gama de Euler, tal que \Gamma(x)=\int_0^\infty e^{-t}t^{x-1}\,dt Assim, \Gamma(x)=g(x-1)\,\! E, analogamente, para números inteiros, \Gamma(n)=(n-1)!\,\! Funções relacionadas [editar] \Gamma(a,x) = \int_x^{\infty} t^{a-1}\,e^{-t}\,dt .\,\! \gamma(a,x) = \int_0^x t^{a-1}\,e^{-t}\,dt .\,\! \psi(x) =\frac{d}{dx} \ln{\Gamma(x)}= \frac{\Gamma'(x)}{\Gamma(x)}. Referências 1. a b c GNU Scientific Library, Reference Manual, 7.19.1 Gamma Functions [em linha] Ícone de esboço Este artigo sobre matemática é um esboço. Você pode ajudar a Wikipédia expandindo-o.
Which Of The Following Is A Healthy Hygiene Practice For Your Nails Nail Fungus – What is Toenail Fungi? If you have actually been questioning what nail fungus appears like, you’re not alone. Having fungal nail infections is a common trouble, but how do you find the fungus as well as get it under control? Thankfully, the good news is that this infection is treatable, and in many cases, you can avoid the requirement for removal. In fact, treating nail fungus at the earliest indicator is crucial to minimizing the risk of permanent damage. Signs and symptoms There are a variety of various sorts of nail fungi, as well as some are much more common than others. Nail fungi is most usual on the huge toe nails, which create at the front as well as sides of the nail. Much less generally, the fungi starts at the base of the nail, which is usually a lot more common in people with a weak body immune system and also serious illnesses. Fungis that infect skin as well as nails are called dermatophytes, though they are sometimes called yeast. Oral drugs are commonly recommended for fungal toe nails. Nevertheless, these medicines can trigger liver damage and can damage your wellness. Oral therapy is generally not as effective as topical therapies, and might take a couple of months to function. Topical medicines may be a much better selection, yet they are less efficient. Along with taking antifungal tablets, you can additionally look for professional treatment for nail fungus. Treatments There are several types of treatments for nail fungus. There are dental drugs like Sporanox, Lamisil, and terbinafine, which have a variety of negative effects as well as can cause liver issues. Dental medications need to be tested in a laboratory to establish their effectiveness. In recent times, applied treatments have actually taken their area. These treatments generally call for a few months before they see any outcomes, so they might be much less reliable for temporary use. Fungal infections usually start on the end of a nail, particularly in the foot. Fungi expands inside the nail as well as can go into cracks or cuts in the skin. The infection can advance right into a bigger issue as the cuticle ends up being broken and enlarged. It can even create the nail to crumble and also discolor. Once this occurs, it is far too late for therapy. It is therefore necessary to obtain instant therapy to stop additional damage. Avoidance Avoidance of nail fungi begins with way of living modifications. You ought to stay clear of choosing your nails, do not go barefoot in public areas, and also prevent letting your shoes splash. Likewise, see to it to put on footwear that fit appropriately, and also prevent sharing your nail files or various other nail-care devices. Put on rubber handwear covers to safeguard your hands when you’re functioning around your home. Likewise, don’t put on other people’s footwear if you’re vulnerable to toenail fungi. Stay clear of limited footwear and socks. These can promote fungal development. Limited shoes can also promote the development of nail fungi. Always use breathable and moisture-wicking socks while wearing shoes. When possible, wear sandals or flip-flops when you remain in public places. Use tidy footwear whenever possible, especially while in the shower. Lastly, make sure to transform your socks routinely and also do not wear the exact same pair of shoes greater than when a week. Reasons For Smelly Feet Smelly feet are a typical trouble for 10% to 15% of the population. The bacteria responsible for this problem, called Kyetococcus sedentarius, produces volatile sulfur compounds that create the feet to scent like rotten eggs. If you believe your feet smell like rotten eggs, it’s time to look for aid. There are a number of reasons for odiferous feet. Continue reading to find one of the most typical reasons. Foot smell If you’re constantly on your feet, after that you have actually most likely seasoned foot smell at one factor or an additional. The human foot contains over 250,000 sweat glands, as well as these gland are fed by bacteria that produce waste as by-products. This waste is what creates foot smell, as well as feet stink more than any other part of the body. To combat foot smell, it is necessary to clean and also dry your feet routinely, clip your nails, and also file off dead skin. Hypothyroidism The signs and symptoms of hypothyroidism might include stinky feet as well as cold hands. Hypothyroidism impacts the thyroid, the gland that manages the body’s temperature. When the hormonal agent is reduced, the body’s metabolic rate slows down. This causes the body to turn to assimilation, which breaks down body tissues for energy. In this phase, muscular tissue strength decreases, causing achy as well as weak feet. Diabetes If you are diabetic, after that you are possibly aware of the aggravation of stinky feet. This condition can make strolling, standing, or resting uncomfortable. Apart from odor, diabetes mellitus can cause troubles with your flow. To reduce the event of these problems, you must utilize a moisturizing lotion or soap on your feet. If you have actually split heels, you must stay clear of using medicated pads or restroom surgical procedure. Moreover, you need to put on shoes that fit properly. If you are not able to discover appropriate footwear, you can consider getting diabetic-specific socks. These socks have additional pillow as well as don’t have flexible tops. They are constructed from fibers that wick away moisture from the skin. Always put on socks when you go to sleep. Never utilize hot pad on your feet! Peripheral neuropathy The treatment of outer neuropathy and also smelly feet depends on the underlying cause. Medicines for diabetes mellitus as well as vitamin shortage are two typical options. If no recognized cause is discovered, treatment might consist of taking over the counter pain relievers or taking an dental medicine. Anti-seizure drugs or topical drugs having capsaicin and lidocaine may be used. If neither of these alternatives work, a physician can suggest a combination of medication to assist the symptoms. Bromodosis If you have foul-smelling feet, you might be experiencing bromodosis. Normally, the problem is caused by inappropriate hygiene, such as seldom cleaning and not altering socks consistently. Microorganisms can expand as well as create the feet to scent, which can be awkward as well as make individuals self-conscious. The bright side is that there are a number of methods to treat the issue. In most cases, house treatments will resolve the problem, yet much more severe situations might need more powerful techniques.
How long does knee bursitis last? The time it takes to heal the condition varies, but results can be achieved in 2 to 8 weeks or less, when a proper swelling management, stretching, and strengthening program is implemented. Will knee bursitis go away? Bursitis often improves over time, so treatment is usually aimed at symptom relief. However, depending on the cause of your knee bursitis and which bursa is infected, your doctor might recommend one or more treatment approaches. Will knee bursitis go away on its own? Often, knee bursitis will get better all by itself as long as it is not caused by an infection. To treat your knee bursitis, you will need to rest the affected joint and protect it from any further harm. Most people feel better within a few weeks with proper treatment. Is it OK to walk with knee bursitis? Inflamed Bursa Treatment Don’t do anything that seems to make your symptoms worse. You can still do low-impact or gentle exercises like a light walk or stationary bike ride. Ice: Put an ice pack on your knee about 3 to 4 times a day. You can also use a bag of frozen veggies like peas or corn. IT IS INTERESTING: Do tendons repair easily? What happens if bursitis is left untreated? Chronic pain: Untreated bursitis can lead to a permanent thickening or enlargement of the bursa, which can cause chronic inflammation and pain. Muscle atrophy: Long term reduced use of joint can lead to decreased physical activity and loss of surrounding muscle. What triggers bursitis? The most common causes of bursitis are injury or overuse, but it can also be caused by infection. Pain, swelling, and tenderness near a joint are the most common signs of bursitis. Bursitis can be treated with rest and medicines to help with the inflammation. Antibiotics are used if infection is found. Should I wear a knee brace for bursitis? There is no quick fix to knee bursitis. Patience combined with treatments recommended by a doctor is the best remedy for bursitis of the knee. A big bulky brace is not needed with knee bursitis, but a comfortable compression knee brace for bursitis or knee sleeve can help reduce swelling of the affected bursa. What is the best exercise for knee bursitis? Heel slide, straight-leg raises and quad sets are good exercises to do if you have knee bursitis. Bursitis is a condition that affects small fluid-filled sacs in your body called bursae. Does knee bursitis show up on xray? Doctors can often diagnose bursitis based on a medical history and physical exam. Testing, if needed, might include: Imaging tests. X-ray images can’t positively establish the diagnosis of bursitis, but they can help to exclude other causes of your discomfort. Should you walk with bursitis? Avoid High-Impact Activities IT IS INTERESTING: Can osteoarthritis be prevented? Running and jumping can make hip pain from arthritis and bursitis worse, so it’s best to avoid them. Walking is a better choice, advises Humphrey. Is walking good for knee pain? You may worry that a walk will put extra pressure on your joints and make the pain worse. But it has the opposite effect. Walking sends more blood and nutrients to your knee joints. This helps them feel better. What cream is good for bursitis? There are also some topical creams you may use for relief, such as Zostrix (capsaicin cream) and Ben Gay-like creams. Is knee bursitis a hard lump? A person suffering from this condition may find it hard to bend the knee and there may be small lumps under the skin of the kneecap. These lumps are actually the thickened bursa tissues that have formed due to the inflammation. What is septic bursitis of the knee? When a bursa is inflamed and infected, it’s called “septic bursitis.” A bursa can become infected when harmful bacteria enter it via a cut, puncture or insect bite. It’s also possible for the infection to come from other parts of the body. In such cases, the origin may be unknown. What happens if a bursa sac ruptures? If the bursitis is left untreated, the fluid filled sack has the potential to rupture. This could then lead to an infection of the surrounding skin. Your podiatrist
Pure Function Signatures Tell All This is a page from my book, Functional Programming, Simplified “In Haskell, a function’s type declaration tells you a whole lot about the function, due to the very strong type system.” Learn You a Haskell for Great Good! One thing you’ll find in FP is that the signatures of pure functions tell you a lot about what those functions do. In fact, it turns out that the signatures of functions in FP applications are much more important than they are in OOP applications. As you’ll see in this lesson: Because pure functions have no side effects, their outputs depend only on their inputs, and all FP values are immutable, pure function signatures tell you exactly what the function does. OOP function signatures When writing OOP applications I never gave much thought to method signatures. When working on development teams I always thought, “Meh, let me see the method source code so I can figure out what it really does.” I remember one time a junior developer wrote what should have been a simple Java “setter” method named setFoo, and its source code looked something like this: public void setFoo(int foo) { this.foo = foo; makeAMeal(foo); foo++; washTheDishes(foo); takeOutTheTrash(); } In reality I don’t remember everything that setter method did, but I clearly remember the foo++ part, and then saw that it the foo and foo++ values in other method calls. A method that —according to its signature — appeared to be a simple setter method was in fact much, much more than that. I hope you can see the problem here: there’s no way to know what’s really happening inside an impure function without looking at its source code. The first moral of this story is that because OOP methods can have side effects, I grew to only trust methods from certain people. The second moral is that this situation can’t happen with pure functions (at least not as blatanly as this). Signatures of pure functions The signatures of pure functions in Scala/FP have much more meaning than OOP functions because:: • They have no side effects • Their output depends only on their inputs • All values are immutable To understand this, let’s play a simple game. A game called, “What can this pure function possible do?” As an example of this — and as a first thought exercise — look at this function signature and ask yourself, “If FOO is a pure function, what can it possibly do?”: def FOO(s: String): Int = ??? Ignore the name FOO; I gave the function a meaningless name so you’d focus only on the rest of the type signature to figure out what this function can possibly do. To solve this problem, let’s walk through some preliminary questions: • Can this function read user input? It can’t have side effects, so, no. • Can it write output to a screen? It can’t have side effects, so, no. • Can it write (or read) information to (or from) a file, database, web service, or any other external data source? No, no, no, and no. So what can it do? If you said that there’s an excellent chance that this function does one of the following things, pat yourself on the back: • Converts a String to an Int • Determines the length of the input string • Calculates a hashcode or checksum for the string Because of the rules of pure functions, those are the only types of things this function can do. Output depends only on input. A second game example Here’s a second example that shows how the signatures of pure functions tell you a lot about what a function does. Given this simple class: case class Person[name: String] What can a pure function with this signature possibly do?: def FOO(people: Seq[Person], n: Int): Person = ??? I’ll pause to let you think about it ... By looking only at the function signature, you can guess that the function probably returns the nth element of the given List[Person]. That’s pretty cool. Because it’s a pure function you know that the Person value that’s returned must be coming from the Seq[Person] that was passed in. Conversely, by removing the n parameter from the function: def FOO(people: Seq[Person]): Person = ??? Can you guess what this function can do? (Pause to let you think ...) My best guesses are: • It’s a head function • It’s a tail function • It’s a Frankenstein’s Monster function that builds one Person from many Persons A third game example Here’s a different variation of the “What can this pure function possibly do?” game. Imagine that you have the beginning of a function signature, where the input parameters are defined, but the return type is undefined: def foo(s: String, i: Int) ... Given only this information, can you answer the “What can this function possibly do?” question? That is, can you answer that question if you don’t know what the function’s return type is? I’ll give you a little space to think about it ... . . . The answer is “no.” Even though foo is a pure function, you can’t tell what it does until you see its return type. But ... Even though you can’t tell exactly what it does, you can guess a little bit. For example, because output depends only on input, these return types are all allowed by the definition of a pure function: def foo1(s: String, i: Int): Char = ??? def foo2(s: String, i: Int): String = ??? def foo3(s: String, i: Int): Int = ??? def foo4(s: String, i: Int): Seq[String] = ??? Even though you can’t tell what this function does without seeing its return type, I find this game fascinating. Where OOP method signatures had no meaning to me, I can make some really good guesses about what FP method signatures are trying to tell me — even when the function name is meaningless. Trying to play the game with an impure method Let’s look at one last example. What can this method possibly do?: def foo(p: Person): Unit = ... Because this method returns Unit (nothing), it can also be written this way: def foo(p: Person) { ... } In either case, what do you think this method can do? Because it doesn’t return anything, it must have a side effect of some sort. You can’t know what those side effects are, but you can guess that it may do any or all of these things: • Write to STDOUT • Write to a file • Write to a database • Write to a web service • Update some other variable(s) with the data in p • Mutate the data in p • Ignore p and do something totally unexpected As you can see, trying to understand what an impure method can possibly do is much more complicated than trying to understand what a pure function can possibly do. As a result of this, I came to understand this phrase: Pure function signatures tell all. Summary As shown in this lesson, when a method has side effects there’s no telling what it does, but when a function is pure its signature lets you make very strong guesses at what it does — even when you can’t see the function name. The features that make this possible are: • The output of a pure function depends only on its inputs • Pure functions have no side effects • All values are immutable What’s next Now that I’ve written several small lessons about pure functions, the next two lessons will show how combining pure functions into applications feels both like (a) algebra and (b) Unix pipelines. books i’ve written
Written by James McDonald May 26, 2023 This is my setup to serve CakePHP from a virtual subdirectory on Windows 11 where http://localhost/subdir/ points to C:/dev/cake_test/webroot/ Run Terminal as Administrator then: Install chocolatey https://chocolatey.org/install Install MySQL, PHP and Nginx using choco choco install mysql choco install mysql.workbench # this one doesn't work at the moment so download and install separately choco install php choco install nginx Make php-cgi a service Download the latest stable version of WinSW https://github.com/winsw/winsw/tree/v3 (I downloaded the WinSW-x64.exe file) Rename it and move it to the PHP dir. The name can be whatever you choose but just make the exe file and the xml file the same name e.g. php-cgi-service.exe php-cgi-service.xml move WinSW-x64.exe C:\tools\php82\php-cgi-service.exe Create a file named php-cgi-service.xml in C:\tools\php82\ <service> <id>php-cgi</id> <name>PHP Fast CGI</name> <description>Run PHP Fast CGI as a Service</description> <!-- <env name="JENKINS_HOME" value="%BASE%"/> --> <executable>C:\tools\php82\php-cgi.exe</executable> <arguments>-b 127.0.0.1:9999</arguments> <log mode="roll"></log> </service> Install the PHP-CGI service # php-cgi-service.exe is the renamed WinSW-x64.exe executeable # you need a file named service.xml in the same directory # as above to be able to install the service php-cgi-service.exe install Edit the Nginx Configuration in C:\tools\nginx-1.25.0\conf\nginx.conf Whenever you make a change restart Nginx with this powershell command: Restart-Service nginx #user nobody; worker_processes 1; #error_log logs/error.log; #error_log logs/error.log notice; #pid logs/nginx.pid; events { worker_connections 1024; } http { include mime.types; default_type application/octet-stream; #log_format main '$remote_addr - $remote_user [$time_local] "$request" ' # '$status $body_bytes_sent "$http_referer" ' # '"$http_user_agent" "$http_x_forwarded_for"'; #access_log logs/access.log main; sendfile on; #tcp_nopush on; #keepalive_timeout 0; keepalive_timeout 65; #gzip on; server { listen 80; server_name localhost; error_log logs/error.log debug; #charset koi8-r; #access_log logs/host.access.log main; location / { root html; index index.html index.htm index.php; } location /subdir/ { index index.php; alias C:/dev/cake/webroot/; # this will serve the file or dir # then the frontend controller (index.php) try_files $uri $uri/ @subdir; location ~ \.php$ { try_files $uri =404; fastcgi_pass 127.0.0.1:9999; fastcgi_split_path_info ^(.+\.php)(/.+)$; include fastcgi_params; fastcgi_intercept_errors on; fastcgi_param PATH_INFO $fastcgi_path_info; fastcgi_param SCRIPT_FILENAME $request_filename; } } location @subdir { # thanks to https://www.paveltashev.com/blog/nginx-how-to-config-domain-subdirectory-to-point-to-a-subfolder/ rewrite ^ /subdir/index.php last; } # this redirects /subdir to /subdir/ location = /subdir { rewrite ^ /subdir/ permanent; } #error_page 404 /404.html; # redirect server error pages to the static page /50x.html # error_page 500 502 503 504 /50x.html; location = /50x.html { root html; } # proxy the PHP scripts to Apache listening on 127.0.0.1:80 # #location ~ \.php$ { # proxy_pass http://127.0.0.1; #} # pass the PHP scripts to FastCGI server listening on 127.0.0.1:9000 # location ~ \.php$ { fastcgi_pass 127.0.0.1:9999; fastcgi_index index.php; fastcgi_intercept_errors on; fastcgi_param SCRIPT_FILENAME $document_root$fastcgi_script_name; include fastcgi_params; try_files $uri =404; } # deny access to .htaccess files, if Apache's document root # concurs with nginx's one # #location ~ /\.ht { # deny all; #} } # another virtual host using mix of IP-, name-, and port-based configuration # #server { # listen 80; # listen 80; # server_name somename alias another.alias; # location / { # root html; # index index.html index.htm; # } #} # HTTPS server # #server { # listen 80 ssl; # server_name localhost; # ssl_certificate cert.pem; # ssl_certificate_key cert.key; # ssl_session_cache shared:SSL:1m; # ssl_session_timeout 5m; # ssl_ciphers HIGH:!aNULL:!MD5; # ssl_prefer_server_ciphers on; # location / { # root html; # index index.html index.htm; # } #} } 0 Comments Submit a Comment Your email address will not be published. Required fields are marked * This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply. The reCAPTCHA verification period has expired. Please reload the page. You May Also Like… Clear HSTS Settings in CHrome Open chrome://net-internals/#hsts enter the domain in the query field and click Query to confirm it has HSTS settings... Ubuntu on Hyper-v It boils town to installing linux-azure # as root or sudo apt-get update apt-get install linux-azure...
Biofeedback behaviour therapy Biofeedback, information supplied instantaneously about an individual’s own physiological processes. Data concerning a person’s cardiovascular activity (blood pressure and heart rate), temperature, brain waves, or muscle tension is monitored electronically and returned, or “fed back,” to that person by a gauge on a meter, a light, or a sound. Though such activity of the autonomic nervous system was once thought to be beyond an individual’s control, it has been shown that an individual can be taught to use the biological data to learn how to voluntarily control the body’s reactions to stress or “outside-the-skin” events. An individual learns through biofeedback training to detect his physical reactions (inside-the-skin events) and establish control over them. Biofeedback training is a type of behaviour therapy that attempts to change learned responses to stressors. It can be very successful in alleviating symptoms (e.g., pain and muscle tension) of a disorder, and its effects can be especially lasting if used in combination with psychotherapy to help the patient understand his reactions to stress. Complaints that have been treated by biofeedback training include migraine headaches, gastrointestinal cramping (e.g., colitis), high blood pressure, tics, and the frequency and severity of epileptic seizures. Theoretically, many psychologists believe it possible to bring under partial control any physiological process that can be continuously monitored and displayed, including electrophysiological activity of the limbic system and other homeostatic processes. Biofeedback training with brain waves has also been useful in enhancing mental functioning. “Alpha (wave) training” elicits the calming and integrative effects of meditation. Theta wave training has led to more focused attention, the control of “mental blocks” during examinations, and the control of anxiety. Learn More in these related articles: Prozac pills. therapeutics: Behavioral therapy ...Rational emotive therapy aims at altering inaccurate or irrational thoughts that lead to negative emotions or maladaptive behaviour. Other behavioral approaches attempt to modify physical responses... Read This Article stress (psychology and biology) ...of diet, such as decreasing intake of alcohol and caffeine. Severe stress may require psychotherapy to uncover and work through the underlying causes. A form of behaviour therapy known as biofeedba... Read This Article in complementary and alternative medicine (CAM) CAM any of various approaches intended to improve or maintain human health that are not part of standard medical care, also known as conventional, or Western, medicine. The various... Read This Article in behaviour therapy The application of experimentally derived principles of learning to the treatment of psychological disorders. The concept derives primarily from work of the Russian psychologist... Read This Article Photograph in communication The exchange of meanings between individuals through a common system of symbols. This article treats the functions, types, and psychology of communication. For a treatment of animal... Read This Article in feedback In biology, a response within a system (molecule, cell, organism, or population) that influences the continued activity or productivity of that system. In essence, it is the control... Read This Article Photograph in homeostasis Homeostasis, any self-regulating process by which biological systems tend to maintain stability. Read This Article Photograph in medicine The practice concerned with the maintenance of health and the prevention, alleviation, or cure of disease. The World Health Organization at its 1978 international conference held... Read This Article in Neal E. Miller American psychologist, who, with John Dollard, developed a theory of motivation based on the satisfaction of psychosocial drives by combining elements of a number of earlier reinforcement... Read This Article × Britannica Kids LEARN MORE Keep Exploring Britannica Margaret Mead education discipline that is concerned with methods of teaching and learning in schools or school-like environments as opposed to various nonformal and informal means of socialization (e.g., rural development projects... Read this Article A close-up view of skin covered with the red raised papules that are characteristic of chickenpox. Diagnose This! Take this Encyclopedia Britannica Heath & Medicine quiz to test your knowledge about symptoms of common illnesses. Take this Quiz View through an endoscope of a polyp, a benign precancerous growth projecting from the inner lining of the colon. cancer group of more than 100 distinct diseases characterized by the uncontrolled growth of abnormal cells in the body. Though cancer has been known since antiquity, some of the most significant advances in... Read this Article Figure 1: The phenomenon of tunneling. Classically, a particle is bound in the central region C if its energy E is less than V0, but in quantum theory the particle may tunnel through the potential barrier and escape. quantum mechanics science dealing with the behaviour of matter and light on the atomic and subatomic scale. It attempts to describe and account for the properties of molecules and atoms and their constituents— electrons,... Read this Article Forensic anthropologist examining a human skull found in a mass grave in Bosnia and Herzegovina, 2005. anthropology “the science of humanity,” which studies human beings in aspects ranging from the biology and evolutionary history of Homo sapiens to the features of society and culture that decisively distinguish humans... Read this Article Shell atomic modelIn the shell atomic model, electrons occupy different energy levels, or shells. The K and L shells are shown for a neon atom. atom smallest unit into which matter can be divided without the release of electrically charged particles. It also is the smallest unit of matter that has the characteristic properties of a chemical element.... Read this Article The SpaceX Dragon capsule being grappled by the International Space Station’s Canadarm2 robotic arm, 2012. 6 Signs It’s Already the Future Sometimes—when watching a good sci-fi movie or stuck in traffic or failing to brew a perfect cup of coffee—we lament the fact that we don’t have futuristic technology now. But future tech may... Read this List Mária Telkes. 10 Women Scientists Who Should Be Famous (or More Famous) Not counting well-known women science Nobelists like Marie Curie or individuals such as Jane Goodall, Rosalind Franklin, and Rachel Carson, whose names appear in textbooks and, from time to time, even... Read this List The visible solar spectrum, ranging from the shortest visible wavelengths (violet light, at 400 nm) to the longest (red light, at 700 nm). Shown in the diagram are prominent Fraunhofer lines, representing wavelengths at which light is absorbed by elements present in the atmosphere of the Sun. light electromagnetic radiation that can be detected by the human eye. Electromagnetic radiation occurs over an extremely wide range of wavelengths, from gamma rays with wavelengths less than about 1 × 10 −11... Read this Article Galen of Pergamum in a lithographic portrait. Doctor Who? Take this Encyclopedia Britannica Health and Medicine quiz to test your knowledge about famous doctors and their contributions to medicine. Take this Quiz Aspirin pills. 7 Drugs that Changed the World People have swallowed elixirs, inhaled vapors, and applied ointments in the name of healing for millennia. But only a small number of substances can be said to have fundamentally revolutionized medicine.... Read this List Hand washing is important in stopping the spread of hand, foot, and mouth disease. Human Health Take this Health Quiz at Enyclopedia Britannica to test your knowledge of various diseases and viruses effecting the human body. Take this Quiz MEDIA FOR: biofeedback Previous Next Citation • MLA • APA • Harvard • Chicago Email You have successfully emailed this. Error when sending the email. Try again later. Edit Mode Biofeedback Behaviour therapy Tips For Editing We welcome suggested improvements to any of our articles. You can make it easier for us to review and, hopefully, publish your contribution by keeping a few points in mind. 1. Encyclopædia Britannica articles are written in a neutral objective tone for a general audience. 2. You may find it helpful to search within the site to see how similar or related subjects are covered. 3. Any text you add should be original, not copied from other sources. 4. At the bottom of the article, feel free to list any sources that support your changes, so that we can fully understand their context. (Internet URLs are the best.) 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( , 1 ). . , , , / , ! . 33 i. # .NET bool Boolean true, false sbyte SByte byte Byte short Intl6 ushort UIntl6 -128 127 0 255 -32 768 32 767 16 0 65 535 16 int Int32 -2 10 2 10 32 uint UInt32 0 4 10 32 long Int64 -9 10 18 9 10 64 ulong UInt64 0 18 10 64 char Char U+0000 U+ffff Unicode 16 float Single 1.5 10" 38 3.4 10 32 double Double 5.0 10 308 1.7 10 15-16 64 decimal Decimal 1.0 10~ 28 7.9 10 28-29 128 string String Unicode object Object 18 18 45 324 28 , 2" 1 . , 1 1 1 [-2"" ,2 ~ - 1], , 1 [0, 2 - 1] (. . 2.5). , , , . , . , . 4 : , 0,381 10 34 2. 381 4, 560,3 10 5603 5 ( ), 0,012 12 , , . double, . i . , , f 1 oat i nt, - . 1 ong doub decimal , . . 2.5, f l o a t } 7 , double 15-16. , ; , ! decimal 28-29 decimal , . . . # .N1 System. , ] , . , : # . , , , <j : double.MaxValue ( System.Doubl.MaxValue) dout uint.MinValue ( System.UInt32.MinValue) u , , , P o s i t i v e l n f i n i t y , Negativelnfinity NaN . ] , . ; , c . , , , . ( ) . i nt, 35 nt, uint, long ulong, . double. 1, 10 i nt ( [ ), 2147483648 ui nt. , , 1. If, 1UL, 1000m ( . 2.3). : , . - 1 # - 1 [ (. 2.2) . - , value types), , [ . , . , (, [). . ( , -, object. . 2 . 2 . # 2.3 . -. . , . , , . 36 2. ( b , b ). , , . . 2 . 3 . , . , . 9. 5 , # , , -. nullable # 2.0 13. , . # . - ( b o x i n g ) , (unboxing). , , : , , . , . , , . , ( !), . 37 , , . , , . , , . . : ( , ) , . , , . . , . , . : , , , , , , . . - : , . , , # , . , , # , - nporpaj . , < . . , , . ] . int ; float ; , . . # , . , i - , ! quantity , , kolich, , , A, tl7_xz prikol. , , ! , 1 ( L) I ( i ) . 39 , . , , , , . I , ! , : a, b = 1; it = 0.1. = 0 . I f : : i nt, ; b i nt, 1; f 1 oat, 1 0.1. , double ( , ), 'loat; 0.1 . , , , : b = 1, = 100: = b * + 25; , , , , . 1 . , , . 1, Z# , . , , . , 0 . , , . . 1 . 40 3. , , , , , < , . . # - : , . . , , i , , : class X // X { int ; // X int ; // X void Y ( ) // { int ; // , - Y int ; // ( ) // ============ 1 ============ int D; int : // // = : // X ( * * ) = this.: // X ( * * * ) // ============ 1 =============== // ============ 2 ============ int D; } } } // D, - ! , // D, - // ============ 2 =============== // // X . X, TJ : , Y. Y . . , , this ( , ** , X, > ( ). , (. , } 41 * * ) . D , , , . , , , , . 3.1 , . 3 . 1 . using System; namespace ConsoleAppl i c a t i o n l { class Classl { static void MainO { int i = 3; double = 4.12; decimal d = 600m; string s = ""; Console.Write( "i = " ) Console.WriteC "y = " ) Console.Write( "d = " ) Console.Write( "s - " ) Consol . Writel_ine( Consol.WriteLine( Console.Writel_ine( Console.Writel_ine( d s ) ) ) ) } } } , Write , WriteLine, . - ( . . 5 9 ) . ( ) . , . . , const, : const int b = 1; const float x = 0:1, = 0 . I f : // const 42 3. , !- . , . , . . 4 n c e j 0 1, ] , . . , , , const int b = 1. = 100: const int = b * + 25: . one , . , . , + 2 , + , 2 . , , . , . # 1 . 3.1 . 3 . 1 . # . 105 . 108, 221 () [] ++ -- . 47 new . 48 typeof . 280 checked . 46 unchecked . 46 . 127 . 47 43 ( ) . 48 . 48 . 48 ++ . 47 -- . 47 () * . 49 . 50 . 50 . 50 ^ ) ( ) . 53 . 54 . 54 < . 54 > . 54 <= . 54 >= . 54 is . 194 as . 194 . 54 1= . 54 & ( ) . 55 . 55 ( ) . 55 1 && . 56 . 56 II ? ; . 56 . 56 = /= 1- += & 44 3. , 3.1 () & < # 2.0 ??, : 13 (. , . 309). ] , . . 3.1 ; , ? . . , b : int = 2, b = 5; + b 7 i nt, = b , ( 5 ) , , . , , . on , . , + b + ( + ) + , = b = = ( = ) . b = , . , F ( i ) + G(i++) H ( i ) F , . , , i ( 1). , ! . 45 1 - [ . , , , . / , , . () , . ; , (). . . , i nt, . , sbyte, byte, short ushort, i nt. , int. [ . 3.1. , , , . , . , , . , . double [ long float decimal ] ulong . 3 . 1 . 46 3. , , i nt, . , float double decimal . int, ui nt long float long double , . . , , , . # , . : (). , , () . . # -, , Excepti on, System. , () , DivideByZeroException, OutOfMemoryException, OverflowException. , Exception. , catch (), . , , . , , . , , . checked unchecked. checked , unchecked . , , , . , : = checked (b + ) ; unchecked { = b + ; // / / 47 , . , , , . , . /checked, . , . ( . . 8 9 ) . # # , , . ( + + ) ( - - ) , , , , . , , , . 3.2 . 3 . 2 . using System; namespace ConsoleApplicationl { class Classl { static void MainO { int x = 3. = 3; Console.Write( " : " ); Consol.WriteLineC ++ ) ; Console.Write( " : " ); Console.WriteLineC ) ; Console.WriteC " : " ); Console.WriteLineC ++ ) ; Console.WriteC " : " ); Console.WriteLineC ) ; 48 3. , : 4 : 4 : 3 : 4 , , , . , ( , 5 7 ) . new new . : new ( [ ] ) , , : object z = new o b j e c t O : int i = new i n t ( ) : II , int i = 0; , , . new ( , ), , , . , . . , OutOfMemoryException. ( -) . int, long, float, double decimal. , ( . . 3.1). . . . , . 49 ( ! ) bool. false, true, true, false. ( ~ ) , , int, uint, long ulong. 3.3. 3 . 3 . BSing System; namespace ConsoleApplicationl { class Ciassl { static void MainO sbyte a = 3, b = -63, = 126; bool d = true; Console.WriteLineC -a ) // -3 Console.WriteLineC -c ) // -126 Console.WriteLineC !d ) // false Console.WriteLineC ~a ) // -4 Console.WriteLineC ~b ) // 62 Console.WriteLineC ~c ) // -127 , , . , . , 1 . : ( ) , , , : long b = 300; int = ( i n t ) b; byte d = (byte) a; // // , . 50 3. , () int, ui nt, long, ulong, float, double decimal. , ( . . 3.1). , int. decimal , 1 System. OverflowExcepti on . . 3.2. , z . , 2 , , 0. NaN ( n o t a number) , . 3 . 2 . +0 -0 +00 00 NaN +Z -z +0 -0 +00 -oo NaN -z +Z -0 +0 -oo +oo NaN +0 +0 -0 +0 -0 NaN NaN NaN -0 -0 +0 -0 +0 NaN NaN NaN +00 +00 -00 NaN NaN +00 -00 NaN 00 00 NaN NaN -00 +oo NaN NaN NaN NaN NaN NaN NaN NaN NaN (I) . int, uint, long, ulong, float, double decimal. , . ( . . 3.1), int. , . , System. Di vi deByZeroExcept i on. , , . 3.3. , z 1 . , . . 34. 51 . , , , 0. 3 . 3 . / +0 -0 -00 +00 00 NaN +Z -z +0 -0 NaN -z +Z -00 +oo -0 +0 NaN +0 +0 -0 NaN NaN +0 -0 NaN -0 -0 +0 NaN NaN -0 +0 NaN +00 +00 +oo 00 NaN NaN NaN 00 +00 oo +oo NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN ( decimal) 0 , 0. (%) - , . , - ( / ) * . , System.DivideByZeroException. , int ( . . 3.1). , - * , , . . 3.4. , z . 3 . 4 . / / +0 -0 +00 00 NaN +Z NaN NaN NaN - X -z -z NaN NaN -x -x NaN +0 +0 +0 NaN NaN +0 +0 NaN -0 -0 -0 NaN NaN -0 -0 NaN +00 NaN NaN NaN NaN NaN NaN NaN -00 NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN NaN ( decimal) , 0. . 3. , 52 , 3.4. 3 . 4 . , using System; namespace Consol eApplicationl { class Classl { s t a t i c void MainO { int x = 11, = 4; float z = 4; Console.WriteLineC z * ) ; // 16 Console.WriteLineC z * le308 ); // " Console.WriteLineC x / ) ; Console. WriteLineC x / z ) ; // 2 Console.WriteLineC x I ) : // 3 Console.WriteLineC le-324 / le-324 ); // NaN // 2.75 } , . 2 / , 2 , . 2 - , , , , 2 / ( ) 2 / / . ( + ) . one int, ui nt, long, ulong, float, double decimal , ( . . 3.1). ] , i nt. decimal , 1 System.OverflowException . 1 . , . 53 . 3.5. , z . , , , 0. 3 . 5 . +0 -0 +00 -oo NaN +00 -00 NaN +0 +0 +0 +00 -00 NaN -0 +0 -0 +00 -oo NaN +00 +00 +00 +00 +00 NaN NaN 00 00 00 NaN -00 NaN NaN NaN NaN NaN NaN NaN NaN ( - ) . int, ui nt, long, ulong, float, double decimal. , ( . . 3.1). , i nt. decimal , System.OverflowException. . 3.6. , z . , . , , - , , 0 , - . 3 . 6 . +0 -0 +00 00 NaN 00 +00 NaN +0 +0 +0 -00 +00 NaN -0 -0 +0 -00 +00 NaN +00 +00 +00 +00 NaN +oo NaN 00 00 00 00 00 NaN NaN NaN NaN NaN NaN NaN NaN NaN 54 3. , ( ) . 1 , . ( ) . ( ) , ( ), ( ). . int, ui nt, long ulong. 3.5. 3 . 5 . using System; namespace ConsoleApplicationl { class Classl { static void MainO byte = 3, b = 9; sbyte = 9, d = -9; Console.WriteLineC a Console.WriteLineC a Console.WriteLineC b Console.WriteLineC Console.WriteLineC d 1 2 1 1 1 ) ) ) ) ) // // // // // 6 12 4 4 -5 ( < , <=, >, >=, ==, ! = ) . , true false. . 3.7 3 . 7 . == true, , fal se != true, , fal se < true, , fal se > true, , fal se <= true, , fal se >= true, , fal se , 5 , int ui nt, 6 , long ulong. 55 , , . . , NaN, , ! = , false ( != true). , ! = ! ( == ) , NaN, <, >, <= >= . , NaN, < false, ! ( >= ) true. : +0 -0 ; - - ; + + . (&, \|, . ( , . . ) . int, ui nt, long ulong. , ( &), 1 , 1. , ( \| ) , 1 , 1. ( 1 , 1. 3.6. 3 . 6 . using System: namespace { ConsoleApplicationl class Classl { static void MainO Console.WriteLineC 6 & 5) Console.WriteLineC 6 \| 5 ) Console.WriteLineC 6 5 ) // 4 // 7 // 3 56 3. , ! (&&) (11) one . true false. . true, ; true. true true. , , . , \ false, false , . 3.7. 3 . 7 . using System; namespace ConsoleApplicationl { class Classl { static void MainO { Console Console Console Console WriteLineC WriteLineC WriteLineC WriteLineC true & & true & & 1 true 1 true J 1 true false true false ); ); ); ); // // // // true false true true } } } (? :) , . _1 ? _2 : , . true, TI , . , . . : , ; , 2 3, , ; 57 , 3 2, , 2; if ( ) . 3.8. 3 . 8 . using System; namespace ConsoleApplicationl { class Classl { s t a t i c void MainO { int a = 11. b = 4; int max = b > a ? b : a; Console.WriteLineC max ); // 11 : , 1, , 1. : i = ( i < n) ? i + 1: 1; , . , ? b : ? d : a T b ^ c d ^ ) . 1 , 2 ( = , +=, -=, - . . ) . . (=): = : , , . , , , . : <- , . 1 , . . , , . 58 3. , ^ = = = = b + / 2: : : 1; = + 0.5: : = = ! , , . . , { . , > . ( = + 0.5). , , . 0.5, ./: , , , . . . , , , : := ; ] - ( . . 4 5 ) . , ] . . ( +=, = , /= . . ) , , . ! , j , += b 6oi = + . , . , i nt. , , , b byte, += b ] : += (byte). . 59 , , , ( = = = ( = ) ) . . , , . . : , . , , # . 11, . - . , , . , 11. # , , . . # Console, System. Write WriteLine . , 3.1. 3.9. 3 . 9 . using System: namespace ConsoleApplicationl { class Classl { s t a t i c void MainO { int double decimal string d s = = = = 3; 4.12: 600m: ""; Console WriteLineC "i - " + i ) ; Console WriteLineC "y = { 0 } \nd - { 1 } " . y. d ) ; Console WriteLineC "s = " + s ) : 1 2 // 3 // // 60 3. , : : 1 = 3 = 4,12 d = 600 s = WriteLine . , Console Write WriteLine . , . , , . Consol , , 3.9 ) . 1 3. . WriteLine , , , , . ) , : , , +. , . # ToStringO. , : Console.WriteLineC "i = " + i.ToStringO ); 2 . WriteLine, . , : , , , ( 2) , : ( ) , ! ( d) . . (. . 146] ( \ ) ( \ t ) . 61 . Console , , . : 1. , , . 2. . Convert, System, Parse, . 3.10 . 3 . 1 0 . using System; namespace ConsoleApplicationl class Classl { static void MainO Console.WriteLineC " string s = Console.ReadLineO: Console.WriteLineC "s = " + s ); ); // 1 Console.WriteLineC " " ); char = (char)Console.ReadO; // 2 Console.ReadLineC); // 3 Console.WriteLineC "c = " + ); string buf; // - Console.WriteLineC " " ); buf = Consol.ReadLineC); int i = Convert.ToInt32( buf ); 1/4 Console.WriteLineC i ) ; Console.WriteLineC " " ); buf = Console.ReadLineO: double x = Convert.ToDouble buf ); // 5 Console.WriteLineC x ) ; Console.WriteLineC " " ); buf = Console.ReadLineO; double - double.ParseC buf ); // // Console.WriteLineC ) ; Console.WriteLineC " " ); buf - Console.ReadLineO; decimal z = decimal.ParseC buf ); // 7 Console.WriteLineC z ) ; } } 62 3. , . 1. , . Read, ( 2 ) . int, , - 1 , (, Enter). int, a char, int char , , ( . . 4 9 ) . 2 3, . , . , , . Enter . Read, ReadLine, , , . 4 5 Convert, 6 7 Parse Double Decimal . N E T , # double decimal. , . , , . , , 1,95-8. , . - , . 2 . . , , , , . 1 , . , . ! 63 , . 11, . 3.11 3.9, , output.txt. , , ...\ConsoleApplication1\bin\Debug. 3 . 1 1 . using System; // 1 using System.10: namespace ConsoleApplicationl { class Classl { static void MainO { StreamWriter f = new StreamWriterC int i = 3; double = 4.12; "output.txt" // 2 decimal d = 600m; string s = ""; f.WriteLineC "i = " + i ); //3 f.WriteLineC "y = { 0 } \nd = { 1 } " . y, d ); //4 f.WriteLineC "s = " + s ): lib f.CloseO; // 6 } } } , : 1. , ( 1 ) . 2. ( 2 ) . 3. - ( 3 - 5 ) . 4. ( 6 ) . , . . . 3.12 , 3.10, input.txt, D:\C#. , . 64 3. , , Visual Studio.NET. File New File... Text File. 3 . 1 2 . using System; using System.10; namespace ConsoleApplicationl { class Classl { s t a t i c void MainO { StreamReader f = new StreamReader( "d:\\C#\\input.txt" ); string s = f.ReadLineC); Console.WriteLineC "s = " + s ); char = (char)f.ReadO; f.ReadLineC); Console.WriteLineC "c = " + ); string buf; buf = f.ReadLineC); int i = Convert.ToInt32( buf ); Console.WriteLineC i ) ; buf = f.ReadLineC); double x = Convert.ToDoubleC buf ); Console.WriteLineC x ) ; buf = f.ReadLineO; double = double.ParseC buf ); Console.WriteLineC ) ; buf = f.ReadLineO; decimal z = decimal.ParseC buf ); Console.WriteLineC z ) ; f.CloseC); Math , . Math, System. : : Sin, Cos, Tan; : ASin, ACos, ATan, ATan2; 65 : Tanh, Sinh, Cosh; : Exp, Log, LoglO; ( ) , , : Abs, Sqrt, Sign; : Ceiling, Floor, Round; , : Mi n, Max; , : Pow, IEEEReminder; : BigMul; : Di vRem. , : . . 3.8. 3 . 8 . Math Abs Acos Asin Atan \|x\| Abs(x) double Acos(double x) double Asin(double x) double Atan(double x) Atan2 double Atan2(double x, double ) , BigMul long BigMul ( i n t , int ) Ceiling double Ceiling(double ) Cos double Cos(double ) Cosh double CoshCdouble ) DivRem DivRem(x, , rem) E. ( ) double 2,71828182845905 Exp double ex () Floor double Floor(double ) IEEERemainder double IEEERemainder(double , double ) Log double \ogex Log(x) , . . & 3. , 66 3.13 Math. . 3 . 1 3 . Math u s i n g System; namespace C o n s o l e A p p l i c a t i o n l { class Classl { s t a t i c void MainO { Console.Write( " x: " ); s t r i n g buf = Console.ReadLine(); d o u b l e x = d o u b l e . P a r s e ( b u f ); Console.WriteLine( Console.Write( " s i n ; " ; " " + Math.Sin(x) ); :" + Math.Max(x, y) ); ); buf = Console.ReadLine(); double = double.Parse Console.WriteLine( b u f ); " } } } 2 = Vtv-' " + 2 t g 2 a + l,610 log10x . 67 , 1 . >i, d o u b l e . 3.14. 1 3 . 1 4 . i n g System: mespace C o n s o l e A p p l i c a t i o n l class { Classl s t a t i c void Main() ( s t r i n g buf: Console.WriteLineC " x" ): buf = Console.ReadLineO: d o u b l e x = C o n v e r t . T o D o u b l e ( b u f ): C o n s o l e . W r i t e L i n e C " a i f a " buf = Console.ReadLineO: d o u b l e a = d o u b l e . P a r s e C b u f ); ); double = Math.Sqrt( Math.PI x ) Math.ExpC 0 . 2 * M a t h . S q r t ( a ) ) + 2 * Math.TanC 2 * a ) + 1.6e3 * M a t h . L o g ] 0 i ' Math.PowCx. 2) ); Console.WriteLineC " x { 0 } a!fa - { ! ) " , x. a }; C o n s o l e . W r i t e L i n e C " = " + ); } } 1, , : # , . , . . 1 , , . . , . . , , . 1 . , 1 , . 68 : , . ( , ). , , - i . , , , , t r u e f a l s e . , b o o l f l a g , bool empty. # , , Camel (. , . 24). , . , , . . . , c o n s t . , , . , 0 1. , . ( , ). , . . . , , . . , . , , , . , . # . # , . , . , 60-70- , , , . , . # - , , . . . . , , . . , , , . 4. , , , . : ( , , ). : 1-; //' * b + ; // f u ! ) ( i. // k ): w h i l e ( t r u e ): / / () , , , . , , . . if switch . if , switch . if if . . 4.1. . 4 . 1 . : if ( ) _1; [ else ; ] 71 " . t r j t , , - . , . e l s e . 1 , , 7. - , , , . , ( ). , , , . : < 0 ) b = 1: < b && ( > d 11 == 0 ) ) b++: e l s e { b = : = 0: ,7 1 } // 2 < b ) if ( < ) m = a: e l s e = : if ( b < ) m = b: e l s e m = : - b > a ) max = b: e l s e max - a: /.' 3 // 4 } 1 e l s e . , , . , . , 2 , < b . , , . . 3 . , e l s e iONiy 1 f . \, 4 ( ), ) , : ; . = b > a ? b : a ; . , 0 < < 1, , . : i f ( 0 < X && < 1 ) . . . 3 , . 4.2. 72 4. . 4 . 2 . , , . , , . 4.1. 4 . 1 . using System; namespace { ConsoleApplicationl class Classl { static void MainO Console.WriteLineC " x" ); string buf = Console.ReadLineO; double x = Convert.ToDoubleC buf ); Console.WriteLineC " " ); buf = Console.ReadLineO: double = double.ParseC buf ); int kol = 0: if ( x x + y * y < l ) k o l = 2 ; else if ( x * x + * < 4 ) kol = 1; Console.WriteLineC " = { 0 } ", kol ); , , if: 73 -f ( temp < 1 ) k o l = 2; T o n s o l e . W r i t e L i n e C - " = { 0 } " , kol ); , i f , , . . : -""oat . ; ... * == b ) ... -f ( Math.AbsCa - b) < i e - 6 ) ... // ! // ! , , . S i n g l e Double E p s i l o n ( , 1.0 + E p s i l o n != 1 . 0 ) . switch s w i t c h ( ) . . 4.3. switch case 1 case 2 case default 1 2 > . 4 . 3 . switch : switch ){ case __1: [ __1 ] case __2: [ __2 ] case : [ ] [ default: ] 74 4. \| . 1 ( c h a r ) . , , . ] . , , d e f a u l t ( ? s w i t c h ). , b r e a k , g o t o r e t u r n : break s w i t c h , f o r , w h i l e do (. break, . 8 4 ) ; g o t o , case s w i t c h ( . g o t o , . 8 3 ) ; r e t u r n , ( return, . 8 7 ) . g o t o , , . 4.2 , . 4 . 2 . u s i n g System; namespace C o n s o l e A p p l i c a t i o n ! : class Classl { s t a t i c void Main() i s t r i n g buf; d o u b l e . b. res, Corisole. Writpi j n e ' " : " ): buf" = C o n s o l e R e a d L i n e C ) : a = double. Parse Console.WriieLineC c h a r op - b u f i; " " ); (char)Console.ReadO: Console.ReaciLmec); C o n s o l e . W r i t e L i n e C " :" buf = Console.ReadLineO; b = d o u b l e . P a r s e C b u f ); ). , , . 75 bool ok = t r u e : s w i t c h (op) I i 1 case ' + : r e s = a + b; b r e a k : 1 case ' : r e s = a - b: b r e a k ; case ' * ' : r e s - a * b: b r e a k : case ' / ' : r e s = a / b: b r e a k : default : r e s = d o u b l e . N a N : ok = f a l s e : b r e a k : i if (ok) Console.WriteLineC ". " + res ): else Console W r i t e L i n e C " " d e f a u l t , , . . switch if , , , . , . , . # : whi 1, repeat, for foreach. . , , . : , (, 4.4). . , , . ^ ( , : . 4.4, ), ( , . 4.4, 6). . , , . , , . 76 4. . 4 . 4 . , , , , . , . . . , 1 . , . , . break, continue, return goto ( . , . 8 3 ) . ( ) . while : while ( ) . , . true, ( ) . , fa 1 se. . , . 77 . false, . , . , , , : 't, = tx, 0 2, <0 " < 10 > 10 , , dX t. . , : . : 1. . 2. , , . 3. . 4. . 5. , 2-4, . 2-4 , . 4.3. . . . , , , . 4 . 3 . , while _5"ng System; -;~iespace Consol eApplicationl class Classl { static void MainO { double Xn = -2, Xk = 12, dX = 2, t = 2. y; Console.WriteLineC "\| x \| \|" ); // double x = Xn; while ( x <= Xk ) { = t; // 1 l i b 1/2 78 4. / 4,3 () f i ( X >= X < 10 ) = t * ; i f ( >= 10 ) = 2 * t; C o n s o l e . W r i t e L i n e C "\| { 0 . 6 } j { 1 . 6 {1.6} += dX: I", x. ): // 11 //' // 2 2 3 4 , ( ( 6 ) . . . , , . , 4. t c k j , . , Ctrl+Break, : w h i l e ( t r u e ) ; - ' . 4.4 ] . 4 . 4 . u s i n g System; namespace C o n s o l 1 i c a t i o n l { class Class! { s t a t i c void MainO string buf: while ( true ) { C o n s o l e . W r i t e L i n e C "1 - _ 1 . 2 - __2. 3 - " ): buf = Console.ReadLineO: switch buf ) // 1 case " 1 " : C o n s o l e . W r i t e L i n e C " - _ 1 " ): 79 break: case "2 // C o n s o l e . W r i t e L i n ene( C "oi;idaxa break; return; case " 3 " default n y i K i a - _ 2 " ), : Console.WriteLineC break; " !" do , . 4.4, , do while ; , , ( b o o l ) . , . , f a l s e - . , , , . , . , , 4.5. 4 . 5 . . s i n g System: -amespace C o n s o l I i c a t i o n i class Classl { s t a t i c void MainO char do answer: C o n s o l e . W r i t e L i n e C " , answer = ( c h a r ) Console.Read ; Console.ReadLineO; w h l i e ( answer != ' ); a?" ): 80 4. , c o s ( x ) = 0,0001. , , 1 . . . , (, ). . , , , , . , . ( 4.6) , . , , , ( ) 2 , . 4 . 6 . using System; namespace { ConsoleApplicationi class Classl { static void MainO double x, l e f t = 0, right = 1; do x = ( l e f t + right ) / 2; if ( ( Math.Cos(x) - x ) * ( Math.Cos(left) - l e f t ) < 0 ) right = x; else l e f t = x; } while ( Math.Abs( right - l e f t ) < le-4 ); Console.WriteLineC " " + x ); 81 for : for ( ; ; ) ; , , . , , : - ( i n t i - 0. j 20: . . . nt k. m; "" ( k = 1 , m = 0 : . . . , , . . b o o l : t r u e , . . . , : -~: ( i n t i = 0 , j = 2 0 ; i < 5 && j > 1 0 ; i + + , j - - ) ... . f o r ( !). 1 100: " t 5 - 0: - V ( i n t i - 1 , i < - 100; i + + ) s += i : 4.7 , 4.3. 4 . 7 . , for . 5 1 n g System: -amespace class { ConsoleApplicationl Classl s t a t i c void MainO d o u b l e Xn = - 2 , Xk = 1 2 . dX = 2. t = 2, y; Console.WriteLineC "\| \|"; f o r ( d o u b l e x = Xn; x <= Xk: x += dX ) - t; i f ( x >= 0 && x < 10 ) = t * x ; // // 1 , 4 , 5 // 2 // 2 & 82 4. 4.7 () i f ( >= 10 ) C o n s o l e . W r i t e L i n e C "\| = 2 * t; {0,6} \| {1.6} \|". , ): / / 2 // 3 } } } , . . , , , 4.3, , w h i l e f o r . , : f o r : f o r ( b l ; 2: ) ; whi 1: : w h i l e ( 2 ) { ; foreach f o r e a c h , , . , , foreach ( . . 136). , . do w h i l e , , , . w h i l e , , , . 83 f o r e a c h . f o r . , , (, 1 ) . . , : , , , ( ); , , ; ( ); , , , , . # , : g o t o ; b r e a k ; c o n t i n u e ; r e t u r n ; t h r o w . , , . . , t h r o w . 93. goto g o t o : goto ; goto case _; goto default; : ; 84 4. goto . , , . , . : (, - ). , goto . , , , . goto switch. goto case _ , goto default default. , # , , . break break , , break. ch ( 6 ) = 10~ 2 = 1+ 2! 4! +... 6! " + +... 2! \| \ < . , . . \| \ > . : , . , . , , . , - , - 85 ( , , ). . break. , , , , , . . , , 1 , , . : , , . : Cn+i C,i ' ^ . +1, : , _ ~ 2\ - " - (2( + 1 ) ) ! (2 + 1)(2 + 2 ) ' 4.8 . 4 . 8 . j s i n g System: 'aniespace Conso 1 eAppl i c a t i o n l { class Class! { s t a t i c void MainO { double e = le-6; const i n t M a x l t e r = 500; C o n s o l e . W r i t e L i n e C " / / :" ); s t r i n g buf = Console.ReadLineO; d o u b l e x = C o n v e r t . T o D o u b l e C b u f ); b o o l done = t r u e ; // d o u b l e ch = 1, = c h : f o r ( i n t n = 0: M a t h . A b s ( c h ) > e; n++ ) { , . 4. 4.8 () c h * = x * x / ( 2 * n l ) / ( 2 * n += c h ; 2 ) ; // if ( n > M a x l t e r ) { done = f a l s e ; break: } if ( done ) C o n s o l e . W r i t e L i n e C " - else " " Console.WriteLineC " + ); ); } } , , . , , d o u b l e , - , . . "!: ( ); ( ) ; ( ); ( ). , , , , ! , , . continue c o n t i n u e , , . 4 . 8 c o n t i n u e : f o r i n t n = 0; M a t h . A b s ( c h ) > e; n++ ) { ch * = x * x / ( 2 * n + 1 ) / ( 2 * n + 2 ) ; += c h ; if ( n <= M a x l t e r ) c o n t i n u e ; done = f a l s e : break; 87 return return . : return [ ]; . void, . , , . , . 11 , , , , . , . . - . # . , , , , . , ( ) . , - . . . 4.5. . 4 . 5 . 88 4. , , , , , (. 4.6). ! 1 j J ;__ j ^nrm J j ! j ! '_ . 4 . 6 . . ( , ), . ; # ( ). , . , , . , , , , . 89 # , ( ), . ( . . 46), . , , , , . . # . # , , Ctrl+C. , . , , . - . . . , , , , , , , . , . , , . , , , , , . , t h r o w . . 4.1 , . System. E x c e p t i o n , , S y s t e m E x c e p t i o n . t r y . 90 4. 4 . 1 . Ari thmeti cExcepti on ( DivideBeZeroException OverFlowException) ArrayTypeMi smatchExcepti on Di vi deByZeroExcepti on FormatException IndexOutOfRangeException InvalidCastException OutOfMemoryExcepti on OverFlowException StackOverR owExcepti on try try : try. . , , ; catch, , ; f i n a l l y , . try: try [ catch ] [ finally ] catch, f i n a l l y , . , . 1. . , , . , try, , . 2. , , . 3. final 1, ( , ). 91 4. , . . . , , , . t r y . c a t c h , . . c a t c h , , . . : catch( ) { ... /* / } catch( ) {.../ / } catch { ... / / } , - , . , . . , , ( ) . : -' { ... // catch ( OvertlowException ) { // O v e r f l o w E x c e p t i o n catch ... DivideByZeroExcepton () .// D i v i d e B y Z e r o E x c e p t i o n ( 0) catch { /7 , . 92 , , finally ( ), , try. , , , , , , . 4.9 , . , . , , , . . 4 . 9 . using System; namespace ConsoleApplicationi { class Program { static void MainO { string buf; double u, i, r; try { Console.WriteLineC " :" ); buf = Console.ReadLineO; u = double.ParseC buf ); Console.WriteLineC " :" ): buf = Console.ReadLineO; r- = double. ParseC buf ); i = u / r; Console.WriteLineC " - " + i ); } catch ( FormatException ) { Console.WriteLineC " !" ); } catch // { Console.WriteLineC " " ); } } } } 93 try . , try catch, , . , , , , . throw. throw , # , . throw , . , System.Exception. . throw , : throw [ ]; catch . , throw, , : :hrow new DivideByZeroExceptionO; throw , 0 new. . , , throw, , catch, . , # : jsing System; class Test static void F ( ) { try { GO; // , } catch ( Exception ) { Console.WriteLineC "Exception in F: " + e.Message ); e = new Exception( "F" ); throw; // 4. } s t a t i c void GO { throw new Exception( "G" ); // } static void MainO try { FO; } catch ( Exception e ) { Console.WriteLineC "Exception in Main: " + e.Message ); F , , Message ( Exception ). . Exception , , , , , : Exception in F: G Exception in Main: G throw : throw e; , , : Exception in F: G Exception in Main: F , . Exception Exception , . . 4.2. 4 . 2 . System.Exception HelpLink U R L Message . . Source 95 STTrace , . " r E x c e p t i o n ~r.rget.Site checked unchecked 3, , , checked unchecked, , , , , : - checked (b + ) ; // ,^checked { // - b + : , . # , , . , . , , . , . , , . , , . . , , . . (, - ) , . 96 4. -, , (, , , , 0 100 1 . . ) . : , , , . : , . , . . - , , . , , , . , . . , switch, i f . for foreach. goto , , switch , , break return. . do , , , . , . . 82. ( , ), . . if , , . true false: bool busy: if ( busy == true ) { ... } if ( busy == false ) { ... } 1 // ! if ( busy ) // ! if ( !busy ) , , . 97 . : if ( < b ) - 1 ; else if ( > b ) = 2; else if ( == b ) = 3: : if ( < b ) = 1: else if ( > b ) = 2: else = 3; : = 3: if ( < b ) = 1 ; if ( > b ) - 2; i f , else : if ( i > 0 ) break: // i <= 3 : if ( z == 0 ) i = j; else i = k; // : i = z == 0 ? j : k; , . . , , switch default, , . 3 . - (), . , . , . , , . -.-: Exception, , , . , , , . , : , , , : , . . . , , . 4. 98 , , !. , , < . , - ( - ) , . , , . , , . , , , - , , . . , . # , 15, , , . , , ). , f . ! , , , . 3-5 , , : s t r i n g but = "qwerty"; d o u b l e ex = 3.1234; int number = 1 2 ; byte if ( done ) C o n s o l e . W r i t e L i n e ! " - else " " if =0; Console.WriteLineC " + ); ): ( x >= 0 && x < 10 ) = t x ; e l s e if ( x >= 10 ) = 2 * t: else = x; , . 99 , , . : II // // { , , , /* / } - // . , : f=a+b; // ! f - + ; , : while ( true ) { while ( < ) { f o r ( i = 0; i < 10; ++i ) { f o r ( j = 0; j < 10; + + j ) { } } } } // // // II // end f o r ( end f o r ( end w h i l e end w h i l e j i ( ( = 0; j < 10; + + j ) = 0; i < 1 0 ; + + i ) x < ) true ) C# , , , , , , . , , 1 . , , [ 2 ] , [ 6 ] . , . 5 : ( . . 13) ( . . 17). , , - Main. , . , , , . , . , , . c l a s s , , - , . , ( ) , : [ ] [ ] class [ : ] - , c l a s s , . # . , . , . , : c l a s s Demo {} . , , 12. , . 101 , . . 5.1. . . . 5 . 1 . new . . , 8 ( . . 175) ") public protected -i internal ( ) ' protected internal ( ) private . , abstract . , 8 ( . . 181) sealed . , 8 ( . . 182) static . 2.0. ( . . 114) 2-6 . , , . , ( ) . : public internal. , , internal. , , , , . 1 . 9. 110 5. : 5.3 () short tl = 3. t2 = 4: i n t = Max( t l , tZ ) : // Console.WriteLineC // : 4 ): i n t z = Max( a + t l . t l / 2 Console.WriteLineC b ): ); // Max Max // : 5 , . . Main . , , , - , , , . . : , . : . . . , . ( ) , . # : -; - ref; o u t : - params. . , -. - , : p u b l i c i n t Calculate( i n t a. r e f i n t b . o u t i n t c , params i n t f ] d ) ... 200 9. 9.1 () { Monster temp = (Monster) obj; if ( this.health > temp.health ) return 1; if ( this.health < temp.health ) return -1; return 0; } string name; int health, ammo; } class CI ass 1 { static void MainO { const int n = 3; Monster[] stado = new Monster[n]; stado[0] = new Monster( 50, 50, "" ); s t a d o [ l ] = new Monster( 80, 80, "" ); stado[2] = new Monster( 40, 10, "" ) ' ; Array.Sort( stado ); // foreach ( Monster el em in stado ) elem.Passport(); } } } : Monster Monster Monster health = 40 ammo = 10 health = 50 ammo = 50 health = 80 ammo = 80 , . . # I Comparer, . ( IComparer) IComparer System. Col lections. CompareTo, , : interface IComparer { int Compare ( object obi, object ob2 ) 201 .NET , , . ( ). ( Name, SortByName) ( Ammo, EortByAmmo) 9.2. , M o n s t e r . 9 . 2 . . s i n g System; . s i n g System.Col l e c t i o n s ; 'amespace C o n s o l e A p p l i c a t i o n i class Monster { p u b l i c Monster( int health, i n t ammo, s t r i n g name ) { this.health = health; this.ammo = ammo; this.name = name; } p u b l i c i n t Ammo { g e t { r e t u r n ammo; set { if ( v a l u e > 0) else ammo = v a l u e ; ammo = 0; } } p u b l i c s t r i n g Name { g e t { r e t u r n name; } virtual public void PassportO { Console.WriteLineC "Monster {0} \t h e a l t h = {1} name, h e a l t h , ammo ); ammo = { 2 } " , } p u b l i c c l a s s SortByName : IComparer // { i n t IComparer.Compare( o b j e c t o b i , o b j e c t ob2 ) 202 9. 9.2 () M o n s t e r ml = ( M o n s t e r ) o b i : M o n s t e r m2 = ( M o n s t e r ) o b 2 ; return String.Compare( p u b l i c c l a s s SortByAmmo int. IComparer.Compare( m1.Name, m2.Name ); IComparer object obi, o b j e c t ob2 { M o n s t e r ml - ( M o n s t e r ) o b i : M o n s t e r ni2 - ( M o n s t e r ) o b 2 ; if ( ml.Ammo > m2.Ammo ) return 1; if ( ml.Ammo < m2.Ammo ) r e t u r n - 1 ; r e t u r n 0. string mt Class { name: health, ammo; Class1 s t a t i c void M a m ( ) const int Monster3 n=3. s t a d o new M o n s t e r ] , s t a d o [ 0 ] = new M o n s t e r ( 50. 5 0 . s t a d o [ l ] = new M o n s t e r ( 8 0 . 8 0 . s t a d o [ 2 ] = new M o n s t e r ( 4 0 . 10. Console.WriteLine( Array.Sort( stado, " "" "" "" ); ); ): : " ); new M o n s t e r SortByName j; f o r e a c h ( M o n s t e r el em in s t a d o ) el em. P a s s p o r t ' , ' ) . Console.WriteLine( " " Array.Sort ( stado. new M o n s t e r . S o r t B y A m m o O f o r e a c h ( M o n s t e r elem in s t a d o : : Monster h e a l t h 50 ammo M o n s t e r h e a l t h = 40 ammo 10 M o n s t e r h e a l t h = 80 ammo 30 50 %: ); ) elem.Passport(); 203 .NET : M o n s t e r hheeaalltthh = 40 ammo 10 M o n s t e r hheeaalltthh = 50 ammo 50 Monster hheeaalltthh = 80 ammo 80 I C o m p a r a b l e , . , . : < >, <= >=, == ! =. , Is. IComparable, fcqual s GetHashCode. o b j e c t . object (. . 183). 9.3 M o n s t e r . h e a l t h , , 9 . 3 . u s i n g System; namespace C o n s o l e A p p l i c a t i o n i c l a s s Monster IComparable { p u b l i c Monster( int health, i n t ammo, s t r i n g name ) { this.health = health; this.ammo = ammo; this.name = name; p u b l i c o v e r r i d e bool Equals( o b j e c t obj { if ( obj == n u l l II GetTypeO != obj.GetType() return false; M o n s t e r temp = ( M o n s t e r ) o b j ; r e t u r n h e a l t h == t e m p . h e a l t h && ammo name public override == temp.ammo == temp.name; i n t GetHashCode() && iP 204 9. 9.3 () { return name.GetHashCode(); } public s t a t i c bool operator == ( Monster a. Monster b ) { return a.Equals( b ); } // // public static bool operator == ( Monster a, Monster b ) // // return ( a.CompareTo( b ) == 0 j; // public s t a t i c bool operator != ( Monster a, Monster b ) { return ! a.Equals( b ); } // // public s t a t i c bool operator != ( Monster a, Monster b ) // // // return ( a.CompareTo( b ) !- 0 ); } public static bool operator < ( Monster a, Monster b ) return ( a.CompareTo( b ) < 0 ); public static bool operator > ( Monster a. Monster b ) return ( a.CompareTo( b ) > 0 ); public s t a t i c bool operator <= ( Monster a. Monster b ) return ( a.CompareTo( b ) <= 0 ); public s t a t i c bool operator >= ( Monster a, Monster b ) return ( a.CompareTo( b ) >= 0 ); public int CompareTo( object obj 205 .NET M o n s t e r temp = ( M o n s t e r ) o b j ; if ( t h i s . h e a l t h > temp.health ) r e t u r n 1; if ( t h i s . h e a l t h < temp.health ) return - 1 : r e t u r n 0; } s t r i n g name; i n t h e a l t h , ammo; } class { Classl s t a t i c void Main() { M o n s t e r = new M o n s t e r ( 7 0 , 8 0 , M o n s t e r = new M o n s t e r ( 8 0 , 8 0 , "" ); "" ); if ( > ) Console.WriteLine( " " e l s e if Console.WriteLine( " " Console.WriteLine( " " else ); ); ); } } : ( ICIoneable) . . , , ( . 9.1, ). , w e m b e r w i s e C l o n e , o b j e c t . , , , (. 9.1, ). . . 9 . 1 . 9. 206 , , , , . . (. 9.1, ). , . , , I C l o n e a b l e 1 9.4 M o n s t e r MemberwiseClone, I C l o n e a b ' r . , MemberwiseClone . , o b j e c t ( p r o t e c t e d ) . 9 . 4 . u s i n g System: namespace ConsoleApplIcationi { c l a s s Monster ICloneable { p u b l i c Monster( i n t h e a l t h , i n t ammo, s t r i n g name ) { this.health = health: this.ammo = ammo: this.name = name; } p u b l i c Monster S h a l l o w C l o n e ( ) / / { return (Monster)this,MemberwiseClone(); } public object CloneO // { r e t u r n new M o n s t e r ( t h i s . h e a l t h , t h i s . a m m o . " " + t h i s . n a m e ); } virtual public void PassportO { Console.WriteLineC "Monster {0} \t h e a l t h = { 1 } name, h e a l t h , ammo ); } s t r i n g name; i n t h e a l t h , ammo; } class { Classl s t a t i c void MainO ammo = { 2 } " , .NET 207 { Monster = new Monster( 70. 80. "" ); Monster X = ; Monster Y = .ShallowClone(); Monster Z = (51).CIone(); } } } X , . , , . Y Z, , . ( I Enumerable) foreach . . N E T , IEnumerable IEnumerator. foreach . , . IEnumerable () GetEnuraerator, IEnumerator (), . IEnumerator : Current, ; MoveNext, ; Reset, . foreach , . , , foreach, : GetEnumerator, Current, MoveNext Reset. , , , , MoveNext 1 , Current . . , , 2.0 , . , . foreach 208 9. , . y i e l d . ( 9.5). , Monster, . . 9 . 5 . using System; using System.Col lections; namespace ConsoleApplicationi { class Monster { . . . } class Daemon { . . . } class Stado ; IEnumerable // 1 { private Monster[] mas; private int n; public StadoO { mas = new Monster[10]; n = 0; } public IEnumerator GetEnumeratoK) { for ( int i = 0; i < n; ++i ) y i e l d return mas[i]; } public void Add( Monster m ) { if ( n >= 10 ) return; mas[n] = m: ++n; class Classl { static void MainO { Stado s = new StadoO; s.Add( new Monster() ); s.Add( new Monster("") ): s.Add( new Daemon ); foreach ( Monster m in s ) m.Passport(); // 2 .NET 209 , 2.0 , , IEnumerable ( 1), ( 2 ) . MoveNext Current IEnumerator. , 9.5, . foreach , . , , , , , , . 9.6 , . 9 . 6 . using System; using System.Collections; namespace ConsoleApplicationi { class Num : IEnumerable { public IEnumerator GetEnumerator() { y i e l d return "one"; y i e l d return "two"; y i e l d return "three"; y i e l d return "oops"; } } class Classl { static void MainO { foreach ( string s in new NumO ) Console.WriteLineC s ); } : one two three oops ( 1 5 ) : using System; using System.Col lections; 111 - , 7 ( . , 154), . - - , - : . o i d ( i n t ) . . , (. . : , , , . , , , , . . , - , , . , , , , , - . . , : -nt X = 1; ; ; y t e = 1: . s h o r t = 1: , , : ; ): ( ) : ( ) : ( 200 ) ; ( / 4 + 1 ) ; - , , , , , return. , , . , - ( . . 4 5 ) . 210 9. namespace { ConsoleApplicationi class Classl { public static IEnumerable Count( int from, int to ) { from = 1; while ( from <= to ) y i e l d return from++: } static void MainO { foreach ( int i in Count( 1, 5 ) ) Console.WriteLineC i ); } } , . 9.7 Stado, 9.5, , Monster ( GetType, object). 9 . 7 . using System; using System.Col lections; using MonsterLib: namespace ConsoleAppl i c a t i o n i { class Monster { . . . } class Daemon { ... } class Stado ; IEnumerable { private Monster[] mas; private int n; public StadoO mas = new Monster[10]; n = 0; public IEnumerator GetEnumeratorO for ( int i = 0 ; i < n; ++i ) y i e l d return mas[i]; public IEnumerable BackwardsO // for ( int i = n - 1; i >= 0; --i ) y i e l d return m a s [ i ] : public IEnumerable MonstersOnlyO // 112 - . -. - r e f : ref , - : v o i d ( ref int ) , , , . , - ( ) . , , , , . - . r e f . , . - - ( 5.4). 5 . 4 . - - u s i n g System; namespace C o n s o l e A p p l i c a t i o n i class Classl { s t a t i c void P( i n t a, ref int b ) { a = 44; b = 33; Console.WriteLine( " {0} {1}", a, b ); " {0} {1}", a, b ); " {0} {!}", a, b ): } s t a t i c void Main() { i n t a = 2 . b = 4; Console.WriteLineC P( a , r e f b ) ; Console.WriteLineC 113 : :;. 2 4 44 33 2 33 , Main , , b , . , , , . , - , , , . , , . , . , , . , , , . : . o u t . , , , . . 5.4 , ( 5.5). 5 . 5 . , s i n g System; "smespace C o n s o l e A p p l i c a t i o n i class Classl { s t a t i c v o i d P( { } a = 44; b = 33: Console.WriteLineC static ^ i n t a. out int b ) " {0} {1}". a, b ); void MainO & 114 5. : 5.5 () int = 2 , ; ( a, out b ) : Console.WriteLineC " { 0 } { ! } " , a, b ); out. , . , . this . , , . this, . this , , , , : '< class Demo double ; public Demo TO // return this; 1 public void Sety( double ) * this. = ; // . new. . : , void. 115 - , , n u l l . "J , , . \| ., , 5.1). , . , ;. 5.6 Demo , ( ) . . 5 . 6 . ..s-ng S y s t e m ; "=mespace C o n s o l e A p p l i c a t i o n i c l a s s Demo { p u b l i c Demo( i n t a, d o u b l e ) // { t h i s . = ; t h i s . = : } p u b l i c double GetyO // { return ; } i n t ; double ; } class { Classl s t a t i c void MainO { Demo a = new Demo( 3 0 0 , 0 . 0 0 2 ) Console.WriteLineC } } a.GetyO ): // // ; 0,002 Demo b = new Demo( 1, 5 . 7 1 ); // Console.WriteLineC // : b.GetyO ); 5,71 116 5. : , . , : class Demo { public DemoC int a ) // 1 { this.a = a: this. = 0.002; } public Demo( double ) // 2 { this.a = 1: t h i s . = ; } Demo x = new Demo( 300 ); Demo = new DemoC 5.71 ); // 1 // 2 . - , -, . this , : class Demo { public DemoC int a ) // 1 { this.a = a; } public DemoC int a, double ) : thisC a ) // 1 { t h i s . = ; } } , , . , # object. , , . 117 < base, . , : p u b l i c DemoC i n t ) : baseC) // 1 { t h i s . = : , . , .'. , . , . , . , . , . , , , . , # ( p r i v a t e ) . . 5.7 , . . 5 . 7 . ( 2.0) . s i n g System; 'STiespace Consol eAppl i c a t i o n i class D { p r i v a t e DC){} s t a t i c DC) / / II a = 200; } static int a: s t a t i c double b = 0.002; public s t a t i c void PrintC) { Console.WriteLineC Console.WriteLineC "a = " + a ); "b = " + b ); 118 5. : 5.7 () class Class2 { static void MainO { D.PrintO; // D d = new DO; // : , ( ), , . 2.0 , static. , , . s t a t i c ( ). , , . 5.8 . 5 . 8 . ( 2.0) using System; namespace ConsoleAppli cati onl { 'ft. s t a t i c class D { s t a t i c int a = 200; s t a t i c double b = 0.002; public static void Print { Console.WriteLine( "a = " + a ); Console.WriteLineC "b = " + b ); class Classl { static void MainO D.PrintO; 119 i , , , . (, , ) . , - ( 5.9) , . 5 . 9 . Monster ' : System; respace ConsoleApplicationi ;i ass Monster public Monster() this.name = "Noname"; this.health = 1 0 0 ; this.ammo = 100; public Monster( string name ) : thisO this.name = name; public Monster( int health, int ammo, string name ) this.name = name; this.health = health; this.ammo = ammo; public int GetName() return name; public int GetHealthO return health; public int GetAmmoO return ammo; public void PassportO 102 5. : , , . new. : Demo b nPW "'.' 11 Demo new Demo\ //' Demo , , (. - . 3 5 ) . , , , . , new OutOfMemoryExcepti on. , . , . , . - , . ), . . , : , , , , , , (. 5.1). ( . . 5.1): 1 1 , . , . , . 120 5. : 5.9 {) Console.WriteLineC "Monster { 0 } \t health = { 1 } ammo = { 2 } ' name, health, ammo ): string name; // int health, ammo; class Classl { s t a t i c void MainO { Monster X = new MonsterO; X.PassportO; Monster Vasia = new MonsterC "Vasia" ); Vasia.PassportO; Monster Masha = new MonsterC 2 0 0 , 2 0 0 , "Masha" ); Masha.PassportO; : "3 J. Monster Noname Monster Vasia health = 100 ammo = 100 health = 100 ammo = 100 Monster Masha health = 200 ammo = 200 (name, health ammo), (GetName, GetHealth, % GetAmmo Passport) , - , . . , . : [ ] [ ] { [ get ] [ set _ ] } . ( public), . 121 , ( g e t ) ( s e t ) . get, set, . set, ( r e a d - o n l y ) , get, ( w r i t e - o n l y ) . # 2.0 get set. , . , , . , publ i , , protected internal, internal, protected private. 2.0 [ ] [ ] _ { [ [ ] [ ] get _ ] [ [ ] [ ] set ] } : j j b l i c class Button: Control private string caption; public string Caption { get { return caption; // , // // } set { // if (caption != value) { caption = value; Button Control Button , Button Control. , , , . , : Button ok = new ButtonO; :k.Caption = "OK"; string s = ok.Caption; // // 122 5. : . . get return, , . set value, . , . , , , . , , , . , , : class private s t a t i c ComplexObject x; public s t a t i c ComplexObject X // // get { if ( == null) { x = new ComplexObject(); // 1- return X; Monster, 5.9, , 3 . Name , 1 , Health Ammo . , . 5 . 1 0 . Monster using System; namespace ConsoleApplication1 class Monster { 1 public Monster() , , readonly. . 123 { this.health = 1 0 0 ; this.ammo = 1 0 0 : this.name = "Noname"; public Monster( string name ) : t h i s O { this.name = name; } public Monster( int health, int ammo, string name ) { this.health = health; this.ammo = ammo; this.name = name; public int Health // Health health { get { return health; } set { if (value > 0) health = value; else health = 0: } public int Ammo // Ammo ammo { get { return ammo; } set { if (value > 0) ammo = value; else ammo = 0; } } public string Name // Name name { get { return name; } ^ 124 5. : 5.10 () public void PassportO Console.WriteLineC"Monster { 0 } \t health = { 1 } ammo = { 2 } ' name, health, ammo); string name: int health, ammo; // } class Classl { static void MainO Monster Masha = new MonsterC 200. 200, "Masha" ); Masha. PassportO; --Masha.Health; // Masha.Ammo += 100; // Masha. PassportO; } } ; Monster Masha health = 200 ammo = 200 Monster Masha health = 199 ammo = 300 , , - ^ , . , - 3\| . , . (pri v a t e ) . - , . , , , . , , 1 . , , 1 , . 125 . . ( ) . , , . , 1-2 : , , . , ( pri vate). , . , . , . . , , . , out. , ref. , , . , . , . , , , . , , , . , ( ), - , . , 4 ( . . 9 5 ) . 6 , , . 'i , , - "5 . , , '* , . . , (). - 3\| , , . , ( ). , . : null . . 6.1 , , int double, . 6.2 . , , 10 100 : int[] w = new i n t [ 1 0 ] ; s t r i n g [ ] z = new s t r i n g [ 1 0 0 ] ; 127 [1] [0] [2] [3] [4] . 6 . 1 . [1] [0] [2] [3] [ ] [4] . 6 . 2 . w i n t [ ] . new 10 , . z s t r i n g [ ] . new 100 , null. , , . () , . , . , int, ui nt, long /long. , : short = . . . ; s t r i n g [ ] z = new string[n + 1 ] ; , (, w 0 9 ) . , : 44] z[i] , . : , IndexOutOfRangeException. 128 6. . , , , : i n t [ ] = new i n t [ 1 0 ] ; i n t [ ] b = ; // b # Array, System. , , , . System.Array ( . . 133). , , , , . # : , (). . : [] [] [] [] [] : = = = = new [ ]; { }; new [] { }; new [ ] { }; , . ( ): int[] int[] b new i n t [ 4 ] ; // 1 // 2 i n t [ ] = { 61. 2, 5, -9 } : i n t [ ] d = new i n t [ ] { 61. 2, 5. -9 II 3 // 4 new i n t [ ] e = new i n t [ 4 ] { 61. 2, 5. -9 // 5 . , , , , . , , , . 1 : , , , , . 129 . : 3-5, . ( 3 ) , . new, ( 2 ) . , , ( 4 ) . , . , , , 6 ( 6.1). 6 . 1 . , " System; -;-espace C o n s o l e A p p l i c a t i o n i class Classl { s t a t i c void MainO { c o n s t i n t n = 6; i n t [ ] a = new i n t [ n ] { 3, 12. 5. -9, C o n s o l e . W r i t e L i n e C " : " ) ; f o r ( i n t i = 0; i < n; ++i ) Console.WriteC " \ t " + a [ i ] ) ; Console.WriteLineC); l o n g sum = 0; // i n t num = 0 ; / / f o r i n t i = 0; i < n; ++i ) i f ( a[i] < 0 ) { sum += a [ i ] ; ++num; } Console.WriteLineC Console.WriteLineC " = " + sum ); "- = " + num ); i n t max = a [ 0 ] ; // f o r ( i n t i = 1 ; i < n; ++i ) i f a [ i ] > max ) max = a [ i ] ; C o n s o l e . W r i t e L i n e C " = " + max ); , . 103 . 5 . 1 . 7J . , , . , . , . (. . 5 7 ) . , , , . , , , (. 5.2). , , a, b , b = . b . , 130 6. . . : [,] [,] [,] [,] [,] ; = = = = new [ _1, _2 ]; { }; new [,] { }; new [ _1, _2 ] { }; ( ): i n t [ , ] a; / I I i n t [ , ] b = new i n t [ 2 . 3 ] ; 111 i n t [ , ] = { { 1 , 2. 3 } . { 4 , 5. 6 } } ; // 3 new i n t [ , ] = new i n t [ . ] { { 1 , 2 , 3 } , { 4 , 5 . 6 } } ; // 4 i n t [ J d = new i n t [ 2 . 3 ] { { 1 . 2 . 3 } , { 4 . 5 , 6 } } ; / / 5 , , , . , , , : [ 1 , 4] b[i. j ] b[j, i] , , . , 3 x 4 (. 6.3). 0 0 /771 1f 01 02 12 13 20 22 23 21 . 6 . 3 . , . ( ) . . . 6.4, 6.2. 131 . 6 . 4 . 6.2 6 . 2 . using System; namespace ConsoleApplication1 class Classl { static void MainO const int m = 3, n = 4; i n t [ , ] a = new int[m, n] { 2,-2. 8. 9 } . {-4,-5. 6,-2 }, & 132 6. {) 6.2 { 7, 0, 1, 1 } }: Console.WriteLineC " :" ); for ( int i = 0; i < m; ++i ) { for ( int j = 0; j < n; ++j ) Console.WriteC "\t" + a [ i , j ] ) ; Console.WriteLineC); } double sum = 0; int nPosEI; for ( int i = 0; i < m; ++i ) { nPosEI = 0; for int j = 0; j < n: ++j ) { sum += a [ i , j ] ; if ( a [ i . j] > 0 ) ++nPosEl; } Console.WriteLineC " {0} { 1 } - ", i , nPosEI ); } Console.WriteLineC " : " + sum / m / n ) ; } } sum . , . , sum , , . . , : , . 133 , . . 6.5. >[0] 1] [1] i [ l ] fo] . 6 . 5 . : [ ] [ ] ; , , , : " = new int[3][]; // [0] = new int[5]; // 0- (5 ) [1] = new int[3]; // 1- (3 ) a[2] = new int[4]; // 2- (4 ) [ 0 ] , [ 1 ] [ 2 ] , ( ) . : \it[][] = { new i n t [ 5 ] , new i n t [ 3 ] , new i n t [ 4 ] }; , , : =:i][2] a[i][j] a[j][i] . , , . System.Array , # A r r a y , , . 6.1. 134 6. 6 . 1 . Array Length ( ) Rank BinarySearch Clear Copy CopyTo GetValue IndexOf LastlndexOf Reverse SetValue Sort Length , , , . . 6.3 Array . 6 . 3 . Array using System; namespace { ConsoleApplicationi class Classl { s t a t i c void MainO { int[] a = { 2 4 , 50, 18, 3, 16, -7, 9, -1 } ; PrintArrayC " :", a ); Console.WriteLineC Array.IndexOf( a, 18 ) ); Array.Sort(a); PrintArray( " :", a ); Console.WriteLineC Array.BinarySearch( a, 18) ); } public s t a t i c void PrintArrayC string header, i n t [ ] a ) 135 { Console.WriteLineC header ) ; for ( int i = 0; i < a.Length; ++i ) Console.WriteC "\t" + a [ i ] ) ; Console.WriteLineC); } } ) Sort, IndexOf BinarySearch , , , . . , , IndexOf. , 18, . ( ) , , , . Classl PrintArray, . : header . Length. , . : / : 24 " -7 50 18 16 -7 -1 16 18 24 50 : -1 PrintArray , , Array. , Get Value, Array . : : . : i i c static void PrintArrayC string header, Array a Console.WriteLineC header ) ; for ( int i = 0; i < a.Length; ++i ) Console.WriteC "\t" + a.GetValue(i) ); Console.WriteLineC); 136 6. 6.4 Array . 6 . 4 . Array using System; namespace { ConsoleApplicationi class Classl / ? { i n t [ ] [ ] a = new i n t [ 3 ] [ ] ; a [ 0 ] = new int [ 5 ] { 24. 50. 18. 3. 16 } ; a [ l ] = new int [ 3 ] { 7, 9. -1 } ; a [ 2 ] = new int [ 4 ] { 6. 15, 3, 1 } ; Console.WriteLineC " :" ); for ( int i = 0; i < a.Length; ++i ) { for ( int j = 0; j < a [ i ] . L e n g t h ; ++j ) Console.WriteC "\t" + a [ i ] [ j ] ); Console.WriteLineC); } Console.WriteLineC Array.IndexOf( a [ 0 ] , 18 ) ); , . : : 24 50 18 -1 15 16 foreach foreach 1 . . , : . : foreach ( in ) 1 , , 9. 137 foreach , ( ) . , , . . , : int[] = { 2 4 , 50, 18, 3, 16, -7, 9, -1 } ; foreach : foreach ( int in ) Console.WriteLineC ); : , . 6.4 foreach , , , for: foreach ( i n t [ ] in ) { foreach ( int in ) Console.WriteC "\t" + ): Console.WriteLineC); } 6.5 , 6.1, foreach. , . 6 . 5 . foreach using System; namespace { ConsoleApplicationi class Classl { s t a t i c void MainO { i n t [ ] a = { 3, 12, 5, -9, 8, -4 } ; Console.WriteLineC " :" ); foreach ( int el em in a ) Console.WriteC "\t" + elem ); Console.WriteLineC); long sum = 0; // int // num = 0 ; foreach int elem in ) if ( elem < 0 ) ^ & 138 6. 6.5 () sum += elem; ++num; } Console.WriteLineC "sum = " + sum ); Console.WriteLineC "num = " + num ); int max = a [ 0 ] ; // foreach ( int elem in a ) if ( elem > max ) max = elem; Console.WriteLineC "max = " + max ); } } } 6.3: public static void PrintArrayC string header, Array a { Console.WriteLineC header ) ; foreach ( object x in a ) Console.WriteC "\t" + x ); Console.WriteLineC); } , , object, , . # ( . . 178), , . foreach , , . , , , . Monster: using System; namespace ConsoleApplicationi { class Monster { . . . } class Classl 139 static void MainO { Random rnd = new Random(); const int n = 5; Monster[] stado = new Monster[n]: // 1 for ( int i = 0; i < n; ++i ) 112 { s t a d o [ i ] = new Monster( rnd.Next( 1, 100 ), rnd.Next( 1 , 200 ) , "Crazy" + i .ToStringO ): } foreach ( Monster x in stado ) x.Passport(); // 3 } } : 'Onster CrazyO 'onster Crazyl health = 18 ammo = 94 health = 85 ammo = 75 ''onster Crazy2 Onster Crazy3 Onster Crazy4 health = 13 ammo = 6 health = 51 ammo = 104 health = 68 ammo = 114 Random, ( . . 148). 1 Monster?, null. 2 : -ew , ( ). 3 foreach . , , . , # - : , , . char Unicode. (. . 26). # Char . N E T System. , , 104 5. : , ( , , b ). . 5 . 2 . . , . , ( b , b nullI). : , , , ( . . 3 8 ) ( . . 4 1 ) . , , . , . : [ ] [ ] [ const ] [ = _ ] , 12, . 5.2. 1-6. 5 . 2 . new public protected internal 140 6. . . 6.2. 6 . 2 . System.Char GetNumericValue , , -1 Unicode- IsControl true, IsDigit true, IsLetter true, IsLetterOrDigit true, IsLower true, IsNumber I1 -sir S GetUnicodeCategory true, ( ) IsPunctuation true, IsSeparator true, Isllpper true, IsWhiteSpace true, ( , ) Parse ) ToLower ToUpper MaxValue, MinValue ( ) 6.6 . 6 . 6 . System.Char using System; namespace { ConsoleApplicationi class Classl { static void MainO { try char b = 1 = 0x63' , d = Ou0032'; // 1 Bee Unicode- , , (DecimalDigitNumber), (LetterNumber), (LineSeparator), (LowercaseLetter) . . 141 Console.WriteLine( " { 0 } { 1 } { 2 } " . b . . d ) ; Console.WriteLine( " { 0 } { 1 } { 2 } " , char.ToLower(b). char.Tol)pper(c), char.GetNumericValue(d) ) ; char a; do // 2 { Console.Write( " : " ); a = char.Parse( Console.ReadLine() ); Console.WriteLine( " { 0 } . - { 1 } " . . int) ) ; if (char.IsLetter(a)) Console.WriteLine(""); if (char. IsUpper(a)) Console.WriteLine("Bepx pe."); i f (char.IsLower(a)) Console.WriteLine(" pe."); i f (char.IsControl ( a ) ) Console.WriteLine("pa"); i f (char.IsNumber(a)) Console.WriteLine(""); i f (char.IsPunctuation(a)) Console.WriteLine("Paee"); } while (a != ' q ' ) : catch Console.WriteLine( " " ); return; 1 . . . 2 . , Ctrl . Parse, , , char. , Enter. , q. 1 . . # , , 2 , , : string s = '' + ' ' + ' ; // - "" , , . . 1 : U n i c o d e ! - ( . . 59). 142 6. , , Array, . 6.1. . 6.7. 6 . 7 . using System; namespace { ConsoleApplicationi class Classl { static void MainO { char[] a = { 'm', 'a', 's', 's', 'i', }; char[] b = " " .ToCharArrayO; PrintArray( " :", ); int pos = Array.IndexOf( a, 'm' ); I T "3 4 a[pos] = ' M ' ; PrintArrayC " a:", a ); if Array.Reverse! b ) ; PrintArrayC PrintArray( " b : " , b ); " b : " , b ); "? # public s t a t i c void PrintArrayC string header, Array a ) Console.WriteLineC header ); foreach ( object x in a ) Console.WriteC x ); Console.WriteLineC "An" ); : : massiv : Massi V : : // 1 // 2 143 , ( 1), ToCharArray string, ( 2 ) . string string, Unicode, # . System.String . N E T . : string string string char[] string s; t = u = = V = "qqq"; new stringO '. 2 0 ) ; { ' 0 ' , ' 0 ' , ' 0 ' }; new string( ); // // // // // 20 (=); ( = = ) ; ( ! = ) ; ( [ ] ) ; () ( + ) . , , . , . , . , string , , . . System.String , , . . 6.3. 6 . 3 . System.String Compare ( ) . . . 144 6. 6.3 () CompareOrdi nal CompareTo Concat Copy Empty ( ) Format ( . ) IndexOf, IndexOf Any, LastlndexOf, LastIndexOfAny Insert Intern, Islnterned , . , Intern , Is Intern nul I Join . ( . ) Length PadLeft, PadRight ( ) Remove Replace Split , . StartsWith, EndsWith true false , Substring ToCharArray ToLower, Tollpper 145 Tim, ( P a d L e f t P a d R i g h t ) rimStart, "rimEnd 6.8. 6 . 8 . string j s i n g System: namespace C o n s o l e A p p l i c a t i o n i class { Classl s t a t i c void MainO { s t r i n g s = " ": C o n s o l e . W r i t e L i n e C s ); s t r i n g sub = s . S u b s t r i n g C 3 ).Remove( 1 2 , 2 ); C o n s o l e . W r i t e L i n e C sub ) ; s t r i n g [ ] mas = s . S p l i t O ' ) ; s t r i n g j o i n e d = s t r i n g . J o m ( "! Console.WriteLineC j o i n e d ); III 112 " , mas ) ; C o n s o l e . W r i t e L i n e C " " ) ; s t r i n g x = Console.ReadLineO; C o n s o l e . W r i t e L i n e C " " + x ); double a = 12.234; i n t b = 29; Console.WriteLineC " a = {0,6;C} b = { 1 , 2 : X } " , a, b ); Console.WriteLineC " a = { 0 , 6 : 0 . # } a = { 1 , 5 : 0 . # ' . a, b ); // 3 II A '}" l i b } : ! ! ! ! ; = 12,23. b = 10 1 : S u b s t r i n g s, , . Remove, , 12-. s u b . 146 6. Split ( 2) , . , mas. Join ( ) mas , "! ". 3 , . 4 6.8 Format, . , , . : { [,[:_]]} . , , . . m , . , . , . , , . , ( C u r r e n c y ) , , X ( H e x a d e c i m a l ) . , . . 5 . , : , #, 0. #, , . 0, , 0. . 6.4 . 6 . 4 . 1,243 00.00 01,24 1,243 #.## 1,24 0,1 00.00 00,10 0,1 #.## ,1 147 , . StringBuilder , string, , . StringBuilder, System.Text . new , : StringBuilder = new StringBuilder( : StringBuilder b = new StringBuilder( "qwerty" StringBuilder = new StringBuilder! 100 ) ; ); StringBuilder d = new StringBuilder! "qwerty", 100 ).; StringBuilder e = new StringBuilder! "qwerty", 1, 3, 100 ); // // // // // : , ( ). , . ( 1), , (16 ). , , / . , 5 3 , ( "wer"). StringBuilder . 6.5. 6 . 5 . System.Text.StringBuilder Append . , , string AppendFormat Capacity . , ArgumentOutOfRangeExcepti on Insert Length ( ) MaxCapacity Remove & 148 6. 6 . 5 () Replace ToString string 6.9. 6 . 9 . StringBuilder using System; using System.Text; namespace ConsoleAppl icationi { class Classl { static void MainO { Console.WriteC " : " ); double salary = double.ParseC Console.ReadLineO ); StringBuilder a = new StringBuiIder(); a.Appendt " " ); a.AppendFormatC " { 0 . 6:C} - { 1 , 6 : C } " . salary, salary 12 ); Console.WriteLineC a ) ; a.ReplaceC " p . " , ".$" ) ; Console.WriteLineC " : " + a ); } } } : : 3500 3 500.00. - 42 000,00. : 3 500,.$ - 42 000..$ , , . Capacity, . Random , , , . # Random, System. 149 < Random , : Ondom = new Random!); Ondom b = new Random! // 111 1 ); : ( 1) , . . i n t ( 2 ) , . , . 6.6. 6 . 6 . System.Random 'iBxt!) i nt '.) [ 0 , ] 8.(, ) [ , ] 'iextByt.es() [ 0 , 2 5 5 ] '.extDoubl() [ 0 . 1) 6.10. 6 . 1 0 . . s i n g System; -amespace C o n s o l e A p p l i c a t i o n i class { Classl s t a t i c void MainO { Random a = new Random(); Random b = new Random( 1 ); c o n s t i n t n = 10; Console.WriteLine! "\n [ 0 , 1 ] : " ) ; f o r ( i n t i = 0; i < n; ++i ) Console.Write! Console.WriteLine! "{0 , 6 : 0 . # # } " , a.NextDouble() ); "An [ 0 , 1 0 0 0 ] : " ) ; f o r ( i n t l = 0; i < n; ++i ) Console.Write! Console.WriteLine! " " + b . N e x t ( 1000 ) ); "\n [ - 1 0 , 1 0 ] : " ) ; 105 protected internal private static readonly volatile ( p r i v a t e ) . , , . . , s t a t i c , , , . , . . ( ) . , . 5.1 Demo . 5 . 1 . Demo, . s i n g System. "amespace Consol eAppl i c a t i o n i class Demo { public public public double i n t a = 1; const double = 1.66; s t a t i c s t r i n g s = "Demo" y; // // II // class Classl { s t a t i c void MainO Demo x = new Demo(); Console.WriteLine! x.a ); C o n s o l e . W r i t e L i n e ! Demo. C o n s o l e . W r i t e L i n e ! Demo.s // Demo ): ); // // // x . a - Demo. - 150 6. 6.10 {) for ( int i = 0; i < n; ++i ) Console.WriteC " " + a.NextC-10, 10) ); Console.WriteLineC "\n [0, 255]:" ); b y t e [ ] mas = new b y t e [ n ] ; a.NextBytesC mas ): for ( i n t i = 0; i < n; + + i ) Console.WriteC " " + mas[i] ); 0,02 [0, 0,4 1]: 0,24 0,55 0,92 0,84 0,9 0,78 0,78 0,74 657 432 354 943 101 642 [0, 1000]: 248 -8 467 [-10, -6 [0, 771 10]: -10 -5 -2 -1 255]: 181 105 60 50 70 77 9 28 133 150 , , , , , . , , . IndexOutOfRangeException, , for. , , . , , . , , , , , , . , , , , , , 13. 151 , # string , . OnngBuilder. - , , . , 15. : , 5. , 5, , . , , , . , , . , . , . , , . (resolution) 1 . . , , , . , , : 1 params, , . 153 // : i n t max( i n t a , i n t b ) // : i n t max( i n t a , i n t b , i n t ) // : i n t max ( i n t a. string b ) // : i n t max ( s t r i n g b, i n t a ) C o n s o l e . W r i t e L i n e C max( 1, 2 ) ); C o n s o l e . W r i t e L i n e C max( 1, 2. 3 ) C o n s o l e . W r i t e L i n e C max( 1, "2" ); ): C o n s o l e . W r i t e L i n e C max( " 1 " , 2 ) ) ; max , ( ). , , , b o o l c h a r i n t , f l o a t d o u b l e . . , . , , , , - ( . . 4 5 ) . , , . , . ( . . 106). , , , ( o u t r e f ) . , , , : i n t maxC i n t a . int b ) i n t max( i n t a , r e f i n t b ) , . , , , . . N E T . , C o n s o l e W r i t e L i n e 19 . , . . , . , 154 7. : , , . . , , . , , . ( , ). , ( - 1 ) . , 0 != 1 1 != 1: long facte long n ) { if ( n == 0 11 n == 1 ) return 1; return ( n * fact( n - 1 ) ); // // } To : long factt long n ) { return ( n > 1 ) ? n * f a c t e n - 1 ) : 1 : } , , , , ( 13). , . , , , . , . # params. , , , : public int Calculate( int a, out int c. params i n t [ ] d ) ... . , , . 155 Length. ( 7.1). 7 . 1 . .sing System; 'smespace ConsoleApplicationi class Classl { public static double Average! params i n t [ ] a ) { if ( a.Length == 0 ) throw new Exception! " " ); double av = 0; foreach ( int elem in a ) av += elem; return av / a.Length; } static void Main!) { try { i n t [ ] a = { 10, 20, 30 } ; Console.WriteLine! Average! a ) ); intC] b = { -11. -4, 12, 14, 32, -1, 28 }; Console.WriteLine! Average! b ) ); short z = 1. e = 12; byte v = 100; Console.WriteLine! Average! z, e, v ) ); Console.WriteLine! Average!) ) ; III /12 //3 //4 } catch! Exception e ) { Console.WriteLine! e.Message ) ; return; } } } : -^ Average . . , (NaN) . 156 7. : - . , . Main , , Main . . : // static MainO { ... } static void MainO { ... } // : static Main( string[] args ) { / * . . . * / } static void Main( string[] args ) { / * . . . * / } , , . args. , args. , , , void. Main , , . , : static int Main( s t r i n g [ ] args { if ( . . . /* / ) return 1; if ( . . . / / ) return 100; } , . , , . 7.2 Main, . 7 . 2 . Main using System; namespace ConsoleApplicationi { class Classl 157 static void Main( s t r i n g [ ] args ) { foreach( string arg in args ) Console.WriteLineC arg ); Console.ReadO; } } ConsoleApplication1.exe : ; cs\ConsoleApplicationl\bin\Debug\ConsoleApplicationl.exe one two three " , . , , . . , , , . , . : this [ _ ] get set , . [ , 12, 158 7. : public, . . , ( g e t ) ( s e t ) . get, set, . set, (read-only), get, ( w r i t e - o n l y ) . , . . , - . 7.3 -, [, 100]. , , . 7 . 3 . using System; namespace ConsoleApplicationi { class SafeArray { public SafeArray( int size ) // a = new i n t [ s i z e ] ; length = size; public int Length // - get { return length; } public int t h i s [ i n t i] // get { if ( i >= 0 && i < length ) return a [ i ] ; else { error = true; return 0; } } set { if ( i >= 0 && i < length && value >= 0 && value <= 100 ) a [ i ] = value; 1 C# 2.0 , ( , . . 120). 159 else error = true; l } public bool error = false; // i n t [ ] a; // int // length; } class Classl { static void MainO { int n = 100; SafeArray sa = new SafeArray( n ); // for ( int i = 0; i < n; ++i ) { s a [ i ] = i 2; III Console.WriteC s a [ i ] ); 112 } if ( sa.error ) Console.WriteC " !" ); 11 , . , , , . ( 1), get. ( 2 ) , set. SafeArray : , ; , 0; error, true. error - , , , ( ). . , . , - . 7.4 Pow2, 2. 106 5. : : , ( Classl). , . (, int 0, nu 1 1 ) . , . , readonly . , ( ). , > , . . , } ' . , , . , . [ ] [ ] _ ( [ ] ) ' . . , , , 1-7 . 5.2, , , virtual, sealed, override, a b s t r a c t extern, . public, - , , ( s t a t i c ) , , , . Main. , . 160 7. : 7 . 4 . using System; namespace ConsoleApplicationi { class Pow2 { public ulong t h i s [ i n t i] { get { i f ( i >= 0 ) { ulong res = 1; for ( int k = 0; k < i; k++ ) unchecked { res = 2; // III return res; } else return 0; } } } class Classl { static void MainO { int n = 13; Pow2 pow2 = new Pow2(); for ( int i = 0; i < n; ++i ) Console.WriteLineC "{0}\t{l}O i, pow2[i] ); 1 , , , . , , Pow2 , ulong, ? . : 0 16 32 64 ( . . 4 6 ) . 161 128 256 512 24 2048 12 4096 # . , . : - t [ , ] : , c o l i c int t h i s [ i n t i , int j ] ? , . , , : "..Object ., . ; : = + ; // MyObject . , , , . , . ( -). . : ] 12, public static. , (extern). operator, . , . , . 162 7. : . , ( ), . ( public s t a t i c ) ; ( ref out); ; , , , ( ). # : , . : + ++ -- true false operator ( ) : public static int operator +( MyObject m ) public static MyObject operator - - ( MyObject m ) public s t a t i c bool operator true( MyObject m ) , , , . : +, -, ! - ; ++ - - , ; true false bool. . , , , , . ( , ). 163 true fal se SQL, , , . 7.3 SafeArray . : , ; Print ; , . 7.5. 7 . 5 . SafeArray using System; namespace ConsoleApplicationi class SafeArray { public SafeArray( int size ) // a = new i n t [ s i z e ] ; length - s i z e ; public SafeArray( params i n t [ ] arr ) // length = arr.Length; a = new i n t D e n g t h ] ; for ( int i = 0; i < length; ++i ) a [ i ] = a r r [ i ] ; public s t a t i c SafeArray operator ++( SafeArray x ) // ++ SafeArray temp = new SafeArray( x.length ): for ( int i = 0; i < x.length; ++i ) temp[i] = + + x . a [ i ] ; return temp; public int this [ i n t i ] get // 164 7. : 7.5 () { if ( i >= 0 && i < length ) return a [ i ] : else throw new IndexOutOfRangeException(); // if ( i >= 0 && i < length ) a [ i ] = value; else throw new IndexOutOfRangeException(); // } set { public void Print( string name ) // { Console.WriteLine( name + " : " ); for ( int i = 0; i < length; ++i ) Console.Write( "\t" + a [ i ] ) ; Console.WriteLine(); } i n t [ ] a; int length; // // class Classl { static void MainO { try { SafeArray al = new SafeArray( 5. 2, al.Print(" 1 " ) ; al++; a l . P r i n t ( " 1" ); -1. 1, -2 ); } catch ( Exception e ) // { Console.WriteLine( e.Message ) ; } } } } : + & = = ! = > < > = < = 165 operator _ ( 2) : C j b l i c static MyObject operator + Cublic static bool ( MyObject ml, MyObject m2 ) operator == ( MyObject ml, MyObject m2 ) , , , . . == ! =, > <, >= <= . , , , . , , 9 ( . . 203). SafeArray, , 7.6. , . 7 . 6 . SafeArray using System; ramespace ConsoleApplicationl class SafeArray { public SafeArray( int size ) a = new i n t [ s i z e ] ; length = s i z e ; public SafeArray( params i n t [ ] arr ) length = arr.Length; a = new i n t [ l e n g t h ] ; for ( int i = 0; i < length: ++i ) a [ i ] = a r r [ i ] ; public static SafeArray operator + ( SafeArray x. SafeArray ) // + int I en = x.length < y.length ? x.length : y.length; SafeArray temp = new SafeArray d e n ) ; for ( int i = 0; i < Ten; ++i ) temp[i] = x [ i ] + y [ i ] : return temp; " 166 7. : 7.6 () public s t a t i c SafeArray operator + ( SafeArray x, int ) // + SafeArray temp = new SafeArrayCx.length); for ( int i = 0; i < x.length: ++i ) temp[i] = x [ i ] + y, return temp; public s t a t i c SafeArray operator + ( int x, SafeArray ) // + SafeArray temp = new SafeArray(y.length); for ( int i = 0; i < y.length; ++i ) temp[i] = x + y [ i ] ; return temp; public static SafeArray operator ++ ( SafeArray x ) // ++ SafeArray temp = new SafeArrayU.length); for ( int i = 0: i < x.length; ++i ) temp[i] = + + x . a [ i j ; return temp; public int t h i s [ i n t i] II [] get { if ( i >= 0 && i < length ) return a [ i ] : else throw new IndexOutOfRangeExceptionC); set { if ( i >= 0 && i < length ) a [ i ] = value; else throw new IndexOutOfRangeExceptionC); public void PrintC string name ) { Console.WriteLineC name + " : " ); for ( int i = 0; i < length; ++i ) Console.WriteC "\t" + a [ i ] Console.WriteLineC); i n t [ ] a; // int length; // class Classl { static void MainO 167 try SafeArray al = new SafeArrayC 5, 2, -1, 1, -2 ); a l . P r i n t ( " 1" ); SafeArray a2 = new SafeArray( 1, 0, 3 ); a2.Print( " 2" ); SafeArray a3 = al + a2; a3.Print( " 1 2" ); al = al + 100; // 1 a l . P r i n t ( " 1 + 100" ); al = 100 + a l ; // 2 al.PrintC "100 + 1" ); a2 += ++a2 + 1 ; 113 a2.Print( "++a2, a2 + a2 + 1" ); catch ( Exception e ) { Console.WriteLine( e.Message ) ; : 1: 5 i 2: -1 1 0 1 2: 7 3 J 1 + 100: 106 103 100 + 1: 3 100 102 99 206 203 ++2. 2 + 2 + 1: 200 202 199 : , , ( 2 1 ) . += ( 3) , . 168 7. : , . 3 , . . : temp.a[i] = + y . a [ i ] . . : implicit operator ( ) explicit operator ( ) // // , . , . , , . 1 . Monster, 5: public static implicit operator i n t ( Monster m ) return m.health: } public static e x p l i c i t operator Monster( int h ) { return new Monster( h. 100, "Fromlnt" ); . , : Monster Masha = new Monster( 200, 200, "Masha" ); int i = Masha; // Masha = (Monster) 500; // , ; , ; , object , . 169 . . , , . implicit e x p l i c i t , , . , . , . # , . . , , , , , , ( , . . ) . : [ ] [ extern ] - , , . ( ~ ) , . , (extern). , . . , , . , , . . , . , . 107 : o u b l i c double GetyO // { return ; } , . , , , 'Stum (. 5.3). , void, return . return .. . 5 . 3 . . , . - . , , Sin Math, , WnteLioe Console ;cuble = 0 . 1 : :; - Math . S i n ( x ) : 2i'isole.Wri t e L i n e ( x ) ; Sin , , Wntel ine . , , , , , . , , , . , 170 7. : . ( ) . , 1 . , Monster Gun. , : using System; namespace { ConsoleApplicationi class Monster { class Gun { } } : , . 9. , , , ( p r i v a t e ) : , , ; , ; , . , , , , , , . (, + -, ). - , , + ++ ( ). , . 1 : (. . 104). 171 , , . , , , . , , , , , , , params, . . , . , . , - . , . , , , . . , , . . , : , , , . # . 173 . , System.Object: [ ] [ ] class [ : ] , , . , , . . . ( . . 120) Monster, . , , Monster, . Monster ( 8.1). 8 . 1 . Daemon, Monster using System; namespace ConsoleApplicationi class Monster class Daemon Monster public Daemon brain = 1: public Daemon( string name, int brain ) : base( name ) // 1 this.brain = brain; public Daemon( int health, int ammo, : base( health, ammo, name ) string name, int brain ) // 2 this.brain = brain; new public void PassportO // 3 & 174 8. 8.1 () { Console.WriteLineC "Daemon { 0 } \t health = { 1 } ammo = { 2 } brain = { 3 } " . Name, Health, Ammo, brain ): } public void ThinkO // 4 { Console.Write( Name + " i s " ); for ( int i = 0; i < brain; ++i ) Console.WriteC " thinking" ); Console.WriteLine( ); } int brain; // } class Classl { static void MainO { Daemon Dima = new Daemon( "Dima", 3 ) // 5 Dima. Passport (); // 6 Dima.ThinkO; // 7 Dima.Health -= 10; // 8 Dima.PassportO; } } Daemon brain Think, , Passport. 1 Monster Daemon . : Daemon Dima Dima is health = 100 ammo = 100 brain = 3 thinking thinking thinking... Daemon Dima health = 90 ammo = 100 brain = 3 , Daemon ( 5 - 7 ) , ( 8) . , 8.1. , . : 1 Monster , Daemon, . 175 , . Daemon. , , , . , , , . , . , ( , 1 2 ) . base. , , base. , , 1 new . 8.1 Passport. . Passport Daemon , . base, : case.PassportO: , . , , , . , . , , Passport, Daemon , : ev public void PassportO base.PassportO; Console.WriteLineC " brain = { 1 } " , brain ); , . (warning) , , ... 176 8. , private, . Passport name, health ammo . , protected, , Monster. . , , . , 1 . , . : , ; , , ; , ; , . , 2 . 8.2 Monster Monster Daemon. 8 . 2 . using System; namespace .ConsoleApplicationl { class Monster { } class Daemon : Monster 1 , . , . .NET 13. , , , , . 177 { . . . / / . 8.1 } class Classl { static void MainO { const int n = 3; Monster[] stado = new Monsterln]; stado[0] = new Monster( "Monia" ); s t a d o [ l ] = new Monster( "Monk" ); stado[2] = new Daemon ( "Dimon", 3 ); foreach ( Monster elem in stado ) elem.PassportO; // 1 for ( int i = 0; i < n; ++i ) stado[i].Ammo = 0 ; Console.WriteLineC); 111 foreach ( Monster elem in stado ) el em. PassportO; // 3 } } } : Monster Monia Monster Monk Monster Dimon Monster Monia Monster Monk Monster Dimon health = 100 ammo = 100 health = 100 ammo = 100 health - 100 ammo = 100 health = 100 ammo = 0 health = 100 ammo = 0 health = 100 ammo = 0 : Daemon , Monster, , . 2, 1 3 , Passport, . , , , . , , , . : , , , ( ). , (, stadoli ] .Ammo). , 178 8. , . , , , , , , . # , . , , , . , , , ', , . , - , -. # virtual. , : virtual public void Passport!) ... virtual . , , , . , , , (Virtual Method Table. V M T ) . ( ) . . V M T . , . V M T , . , override, : override public void PassportO ... , . , , , , . 179 8.2 virtual override Passport, , ( 8.3). 8 . 3 . using System; namespace ConsoleApplicationi class Monster { virtual public void PassportO { Console.WriteLineC "Monster { 0 } \t health = { 1 } ammo = { 2 } " , name, health, ammo ); } } class Daemon ; Monster { override public void PassportO { Console.WriteLineC "Daemon {0} \t health = { 1 } ammo = { 2 } brain = { 3 } " , Name. Health, Ammo, brain ); } } class Classl { static void MainO { const int n = 3; Monster[] stado = new Monster[n]; stado[0] = new MonsterC "Monia" ); s t a d o [ l ] = new MonsterC "Monk" ): stado[2] = new Daemon ( "Dimon", 3 ); foreach ( Monster elem in stado ) elem.PassportO; for ( int i = 0; i < n; ++i ) stado[i].Ammo = 0; Console.WriteLineC): foreach ( Monster elem in stado ) elem.PassportO; } } } 108 . . . , S i n : p u b l i c s t a t i c double Sin( double a ): , , . . 5.2 Demo ( , , ) . , s , p r i v a t e , Gets, . 5 . 2 . using System; namespace ConsoleApplicationl { class Demo i pub'ic int = 1 ; p u b l i c c o n s t d o u b l e - 1.66; s t a t i c s t r i n g s = "Demo"; double y; p u b l i c double GetyO // { return ; } p u b l i c v o i d SetyC d o u b l e y_ ) // { = _: } p u b l i c s t a t i c s t r i n g Gets // s { return s; class Classl { s t a t i c void MainO { Demo x = new Demo(); x.Sety(0.12): // 180 8. : Monster Monia health = 100 ammo = 100 health = 100 ammo = 100 health = 100 ammo = 100 brain = 3 Monster Monk Daemon Dimon Monster Monia health = 100 ammo = 0 Monster Monk health = 100 ammo = 0 Daemon Dimon health = 100 ammo = 0 brain = 3 , 1 3 Passport, , . . : , . : V M T , V M T , . , , . , , V M T , . - . , . . . , . , , , . , -. , . 181 . , , . . . , , . , . , . abstract. , , . . , , : abstract class S p i r i t public abstract void PassportO: } class Monster Spirit { override public void PassportO { Console.Writel_ine( "Monster { 0 } \t health = { 1 } ammo = { 2 } " , name, health, ammo ); } } class Daemon : Monster { override public void PassportO { Console. Writel_ine( "Daemon { 0 } \t health = { 1 } ammo = { 2 } brain = { 3 } " , Name, Health, Ammo, brain ); } . . . // 12 8. 182 , , . , , , . , . . . . foreach (. . 136) , P r i n t A r r a y . 10. # s e a l e d , , , , : sealed class Spirit } // c l a s s Monster : Spirit { ... s e a l e d . , , , : . , , , . , , , , . , , . , , . . 8.4. 8 . 4 . - u s i n g System; namespace ConsoleApplicationi { class { p u b l i c v o i d ) 183 object { Console.WriteLine( "!!" ); } class { public () { = new ; = new ; } public void _() { .(); .); } , ; } class Classl { static void MainO { AH24_1 = new ); 24_1.3_(); } } : !! !! _ , , , . , (is ) , - (has ). , , , . object System.Object . N E T , # object, . . 184 8. object , . System.Object . Equals true, . : public virtual bool Equals( object obj ); Equals true, . : public static bool Equals( object obi, object ob2 ); GetHashCode - , . . Equals, GetHashCode. - ( . . 291). : public virtual int GetHashCodeO; GetType , , , . . , 1 . : public Type Get ; ReferenceEquals true, . : public static bool( object obi, object ob2 ); ToString , , . , . : public virtual string ToStringO . , Equals , - ( ) , ( ). object Monster 8.5. 1 12 (. . 279). 185 object 8 . 5 . object using System; namespace ConsoleApplicationl class Monster { public Monster( int health, int ammo, string name ) { this.health = health; this.ammo = ammo; this.name = name; } public override bool Equals( object obj ) { if ( obj == null \|\| GetTypeO != obj.GetTypeC) ) return false; Monster temp = (Monster) obj; return health == temp.health && ammo == temp.ammo name == temp.name; && } public override int GetHashCode() { return name.GetHashCode(); } public override string ToStringO { return string.Format( "Monster { 0 } \t health = { 1 } ammo = { 2 } " , name, health, ammo ); } string name; int health, ammo; } class Classl { static void Main() { Monster X = new Monster( 80, 80, "" ); Monster Y = new Monster( 80, 80, "" ); Monster Z = X; if ( X == Y ) Console.WriteLine( X == Y " ) ; else Console.WriteLine( X != Y " ) ; & 186 8. 8.5 () if ( X == Z ) Console.WriteLineC X == Z " ) ; else Console.WriteLineC X ! = Z " ) ; if ( X.Equals(Y) ) Console.WriteLineC " X Equals Y " ); else Console.WriteLineC " X not Equals Y " ); Console.Wri teLi ne(X.GetTypeC)); } } } : X != Y X * Z X Equals Y ConsoleAppli cati onl.Monster Equals . nul 1 , , false. true . GetHashCode . ToString , . , , , Equals . , ( . . 203). . . Y X , Y X, , . , Y . , X, Y, (, ) . , 1 , , . 187 , . , , , . , , , , . , , . , . , , . , , . , , , -. ( ) , , new, , . , , , , ( ) . . , . Y X Y X. , Y X ( Y X ) . , , X Y , Y X, Y X ( Y X ) . , . . , , 1 . , , . , . -. : - . : [ ] [ ] interface _ [ : ] [ ; ] , new, publ i , protected, internal private. new , . . . , ( i n t e r n a l ) . 1 , , 10. 189 , . , , . , , , , , , . I Act ion, , . , , : interface IAction { void Draw(); int Attack(int a ) ; void D i e ( ) ; int Power { get; } } IAction Power, . , , set, : int Power { get; set; } , - . - , , . , , ( Health, Ammo Name ). . publ i , ; 109 Console.WriteLineC x . G e t y O ); C o n s o l e . W r i t e L i n e t Demo.Gets ) // Console.WriteLinet GetsO // ); ); // s // } } } , . , s t a t i c , . , , / . , . : 1. , . 2. . 3. ( ) . 4. . 5. , ; v o i d , , . . . 5.3 . 5 . 3 . . s i n g System; -amespace C o n s o l e A p p l i c a t i o n i class Classl { s t a t i c i n t MaxCint a , i n t b) / / { if ( > b ) r e t u r n ; else r e t u r n b; } s t a t i c void MainO { i n t a = 2, b = 4 : i n t x = Max( a, b ); C o n s o l e . W r i t e L i n e C x ); // Max // : 4 190 9. , , , ( ); , . .NET , . , , IComparable. . , , , . , # . , . , IAction Monster : using System: namespace ConsoleAppl i c a t i o n l { interface IAction { void DrawO; int Attack( int a ) : void D i e O : int Power { g e t ; } } class Monster : IAction { public void DrawO { Console.WriteLineC " " + name ): } public int Attack( int ammo_ ) { ammo -= ammo_; if ( ammo > 0 ) Console.WriteLineC "-!" ); else ammo = 0: 191 r e t u r n ; } public void OieO { Console.WriteLineC " M o n s t e r " + name + " RIP" ); h e a l t h = 0: } public int Power I ciet { r e t u r n ammo * h e a l t h ; } , . publ i c . , 1: Monster V a s i a = new MonsterC 5 0 . 5 0 , "" ); Vasia.DrawC): // M o n s t e r // : I A r t i o n A c t o r = new MonsterC 10, 10, "" ); Actor.DrawC); // // : I Action , . , . , : s t a t i c void Act IAction A ) { A.DrawC); ) s t a t i c v o i d Maine) { M o n s t e r V a s i a = new MonsterC 5 0 , 5 0 , ActC V a s i a "" ); ); , . , , , . 192 9. : . . , : class Monster IAction { int IAction.Power { get { return ammo * health; } } void IAction.DrawO { Console.WriteLineC " " + name ); } } IAction Actor = new Monster( 10, 10, "Mama" ); Actor.Draw(); // // Monster Vasia = new MonsterC 50, 50, "" ); // Vasia.DrawO; , . , - . , , 1 . , , Monster : , : interface ITest { void DrawO; } interface IAction 1 , , . 193 void Draw(); int Attack( int a ) ; void D i e ( ) ; int Power { get; lass Monster : IAction, ITest void ITest.DrawO { Console.Writel_ine( "Testing " + name ); void IAction.DrawO { Console.WriteLineC " " + name ); Draw . . , , : Monster Vasia = new MonsterC 50, 50, "" ); :(ITest)Vasia).DrawO; // : ;(IAction)Vasia).DrawO; //: Testing , , ( public), : slass Monster : IAction. ITest public void DrawO { Console.WriteLineC " " + name ): Draw, , : Monster, IAction ITest. , , . . 9. 1 94 . is as , . is. , , is, , . true, , false . : 1f ( is ) { // "" /'/ "" , - object. , , . , , : static vo'd Act( object A ) { if ( A is IAction ) { IAction Actor = ( I A c t i o n ) A; Actor.Draw(); 1 } Act , , IAction. is , : . as. , , null, : static void Act( object A ) I IAction Actor = A as IAction; if ( Actor != null ) Actor.Draw(); , . 195 -, , . , . , , private internal, (publ i c ) '. , , . - , . new, . . # : interface 1 Base i void F( int i ): } interface I l e f t IBase { new void F( int i interface I n g h t ); // F IBase void G ( ) : interface Iderived : I Left, I Right {} class A void Test( IDerived d ) { d.F( 1 ); // I Left F ( ( I B a s e ) d ) . F ( 1 ): // IBase.F ( ( I L e f t ) d ) . F( 1 ); //' I Left. F v ( I R i g h t ) d ) . F ( 1 ); /7 IBase.F F IBase ILeft, IDerived IRight IBase . , , , . , , . 196 9. , , : class : IRight { IRight.GO { . . . } IBase.F( int i ) { . . . } // IRight.F( int i ) - } , , , : class : { // IRight.GO { . . . } } class : A. IRight { IRight.GO { ... } IBase.F( int i ) { ... } II // } , , . new, . , : interface IBase { void : } class Base : IBase { public void AO { . . . } } class Derived: Base { new public void AO { . . . } } Derived d = new Derived ( ) ; d.AO; IBase id = d; // Derived.AO; id.AO; // Base.AO; , ( ) : 197 interface IBase { void ; } class Base : IBase { public virtual void AO { ... } class Derived: Base { public override void AO { . . . } } Derived d = new Derived ( ) : d.AO; IBase id = d; id.AO; // Derived.AO; // Derived.AO; , , . : , , . IBase _, Base: interface IBase { void A O ; class Base : IBase { void IBase.AO { A _ ( ) ; } protected virtual void A _ ( ) { . . . } class Derived: Base protected override void A _ ( ) { . . . } , -, . : interface IBase { void ; 9. 1 98 } c l a s s Base : IBase I void IBase.A() ... c l a s s D e r i v e d : Base, i i p u b l i c v o i d AO / / D e r i v e d IBase ... , , , : interface Interfacel void FO: } class Classl p u b l i c void F() ... p u b 1 ic v o i d GO ... c l a s s Class-2 Classl. Interfacel new p u b l i c v o i d GO ... j Class2 C l a s s l F. I n t e r f a c e l F. , Class2 , . . , , . .NET ."NET , . , I C o m p a r a b l e , . l E n u m e r a b l e e n u m e r a t e f o r e a c h , I C l o n e a b l e . 199 . , f o r e a c h , A r r a y I E n u m e r a b l e I E n u m e r a t o r . , , . ( IComparable) I C o m p a r a b l e System. CompareTo, : interface IComparable i n t CompareTo( o b j e c t o b j 0, ; , ; , . IComparable M o n s t e r . h e a l t h . 9.1 , , ( , , ). 9 . 1 . IComparable j s i n g System; 'iamespace Consol eAppl i c a t i o n i c l a s s Monster IComparable { p u b l i c MonsterC i n t . h e a l t h , i n t ammo, s t r i n g name ) { this.health = health; this.ammo = ammo: this.name = name; } virtual public void PassportO { Console.WriteLineC " M o n s t e r { 0 } \t h e a l t h = { 1 } name, h e a l t h , ammo ); ammo = { 2 } " , } p u b l i c i n t CompareToC o b j e c t o b j // 211 .NET { for ( int i = 0; i < n; ++i ) if ( mas[i] .GetTypeO .Name == "Monster" ) y i e l d return m a s [ i ] ; } public void Add( Monster m ) { if ( n >= 10 ) return: mas[n] = m; ++n: class Classl { static void MainO { Stado s = new StadoO; s.Add( new Monster() ); s.Add( new Monster("Bac") ): s.Add( new Daemon() ); foreach ( Monster i in s ) foreach ( Monster i in s.Backwards i.PassportO ) foreach ( Monster i in s.MonstersOnly() i.Passport ) i.PassportO } , , . , get , 1 IEnumerable IEnumerator . : y i e l d return , ; y i e l d break . y i e l d . , . -, MoveNext , y i e l d . MoveNext - , . IEnumerable<T> IEnumerator<T> System. C o l l e c t i o n s .Generic, 13. 9. 212 -\ . , ^ ^ ^ , : ^ ^\ crnvi Q , a -fte , .\ \ , , , ; , ; , ( ); , # . : , , . ! ! , , ! , , . [ ] [ ] struct [ : ] _ [ ; ] , , public, internal private ( ; ). , , . , , , , , , , . , : , : , ' protected protected internal; (abstract), ( s e a l e d ) ; 213 ; ( o v e r r i d e ) , o b j e c t ; t h i s , , ; , ( , n u l 1 ) . 9.8 , . . T o S t r i n g : , C o n s o l e . W r i t e L i n e . . 9 . 8 . u s i n g System; namespace ConsoleApplicati onl i s t r u c t Complex { p u b l i c double r e , im; p u b l i c Complex( d o u b l e r e _ , d o u b l e i m _ ) re = r e _ ; im = i m _ ; / / t h i s . r e , p u b l i c s t a t i c Complex o p e r a t o r + ( Complex a. r e t u r n new Complex( a.re + b.re, this.im Complex b ) a . i m + b . i m ); public override string ToStringO r e t u r n ( s t r i n g . F o r m a t ( " ( { 0 . 2 : 0 . # # } ; { 1 , 2 : 0 . # # } ) " . r e , im ) ); } class { Classl s t a t i c void MainO { Complex a = new Complex! 1.2345, 5 . 6 ); 214 9. 9.8 () Console.WriteLineC " = " + ); Complex b: b.re = 10; b.im = 1; Console.WriteLineC "b = " + b ); Complex = new ComplexO; Console.WriteLineC "c = " + ); = a + b; Console.WriteLineC "c = " + ); } } } : = = = = (1.23:5,6) (10; 1) ( 0; 0) (11,23:6.6) , . ( , . . 3 6 ) , , , , . , , , . . ref out. , . , 100 101 , . , : Complex [] mas = new Complex[4]; for ( int i = 0; i < 4; ++i ) { mas[i].re = i; mas[i].im = 2 * i; } foreach ( Complex elem in mas ) Console.WriteLine( elem ); 215 Main 9.5, : ( ( ( ( 0 1 2 3 0) 2) 4) 6) , . , , : enum Menu { Read, W r i t e , Append, E x i t } enum { , , , . i n t , 0, , : enum Nums { two = 2, t h r e e , f o u r , t e n = 10, e l e v e n , f i f t y = t e n + 40 }: t h r e e f o u r 3 4, e l e v e n 11. , . , ; , , , . : [ ] [ ] enum _ [ : ] _ [ ; ] , , new, p u b l i c , p r o t e c t e d , i n t e r n a l p r i v a t e . , . i n t , , ( c h a r ) , : b y t e , s b y t e , s h o r t , u s h o r t , i n t , u i n t , l o n g u l o n g . , i n t . , . , . p u b l i c . 216 9. , - . 9.9. 9 . 9 . using System; namespace ConsoleApplicationl { struct { public enum _ { , , , } public string ; public _3 ; } class Classl { static void MainO { x; x. = ""; x. = ._.; Console.WriteLineC . + " " + . ); } } } : , : enum Flags ; byte { , . 2, = 0x04, 4 = 0x08, 5 = 0x10, 6 = 0x20, 7 = 0x40 ( + , -, ++, - - ) , ( * , &, \|, ~ ) , ( < , <=, >, >=, ==, ! = ) ( s i z e o f ) . . , 217 , , . . : Flags = Flags.2 \| Flags.4; Console.WriteLineC " = {0} { 0 , 2 : } " , ); ++; Console.WriteLineC " = {0} int = ( i n t ) ; { 0 . 2 : X } " . ): Console.WriteLine( "x = {0} { 0 . 2 : X } " , x ); Flags b = (Flags) 65; Console.WriteLine( "b = {0} { 0 . 2 : X } " , b ); ( { 0 , 2 : } ): = 10 = 11 = 11 D = 65 0 0 41 9.10. 9 . 1 0 . using System; namespace ConsoleApplicationi { struct { public enum _ { , , , , , } public string ; public _ ; } class Classl { static void MainO { x; x. = " x. = ._.; for ( int i = 1976; i < 2006; i += 5 ) { if ( x. < ._. ) ++.; 218 9. 9.10 () Console.WriteLineC " : {0} {1} { 2 } " . 1 . . . . 1976 1981 1986 1991 1996 2001 System.Enum # System.Enum, . GetName , : Console.WriteLineC Enum.GetName(typeof( Flags ) . 8 ) ) ; // fc>4 Console.WriteLineC Enum.GetNameCtypeof( ._ ) , ! ) ) : // typeof (. 12). GetNames GetValues , , , , : Array names = Enum.GetNames( typeof(Flags) ); Console.WriteLineC " : " + names.Length ); foreach ( string elem in names ) Console.WriteC " " + elem ); Array values = Enum.GetValuesC typeof(Flags) ); foreach ( Flags elem in values ) Console.WriteC " " + (byte) elem ); IsDefined true, , false , : if ( Enum.IsDefinedC typeof( Flags ), "5" ) ) Console.WriteLine( " 5 " ); else Console.WriteLine( " 5 " ); GetUnderlyingType , . , Flags System.Byte: Console.WriteLine( Enum.GetUnderlyingType( typeof(Flags) ) ); 219 . , , , , -. - , , . # . . , . . N E T . . , IComparable IComparer. IEnumerable IEnumerator foreach, ICloneable . . , , , . , . , , . , , , . 10 , # , . . , . , , , \ . , . . , < : [ ] [ ] delegate ( [ ] ) , , new, public, protected, internal private. , , ! , . 10 , # , . . , . , , , \ . , . . , < : [ ] [ ] delegate ( [ ] ) , , new, public, protected, internal private. , , ! , . 221 : public delegate void D ( int i ); , , void . , , . System.Delegate, , . , . , , . . : , ; , ; . . 10.1 , (001 Hack). 1 0 . 1 . using System: namespace ConsoleApplicationi delegate void Del ( ref string s ); // class Classl public s t a t i c void C001 ( ref string s ) // 1 string temp = " " ; for ( int i = 0: i < s.Length: ++i ) 222 10. , 10.1 () if ( s [ i ] == ' ' 11 s [ i ] ' 0 ' ) temp += ' O ' ; else i f ( s [ i ] = = ' I ' ) temp + = ' ; else temp += s [ i j ; } s = temp; } public static void Hack ( ref string s ) // 2 { string temp = " " ; for ( int i = 0; i < s.Length; ++i ) if ( i / 2 * 2 == i ) temp += char.ToUpper( s [ i ] ); else temp += s [ i ] ; s = temp; } static void MainO { string s = "cool Del d; hackers"; // for ( int i = 0; i < 2; ++i ) { d = new Del( COOl ); // 1 if ( i == 1 ) d = new Del(Hack); // 2 d( ref s ); // Console.WriteLine( s ) ; } : cOOl hackers COOl hAcKeRs , . , , . Combine, System.Delegate, , , . Main , : 223 static void MainO string s = "cool hackers"; Del d = new Del ( C001 ) ; d += new Del( Hack ); // d( ref s ) ; Console.WriteLineC s ); // : C001 hAcKeRs , ; , ; out , , ( , voi d); , , , ; , , System. Null Ref erenceExcepti on. . . , . , . , , , , . . , , . , , . 224 10. , . , . (observer) , . ( 10.2), . O O P S ! . . 1 0 . 2 . using System; namespace ConsoleApplicationi { public delegate void Del( object ); class Subj Del dels: // // - // public void Register( Del d ) // dels += d: public void OOPS // - Console.WriteLineC "OOPS!" ) : if ( dels != null ) dels( this ); // class ObsA public void Do( object ) // - // Console.WriteLineC ", OOPS!" ): class ObsB public static void SeeC object ) // - // Console.WriteLineC " , OOPS!" ): 225 \ :lass Classl { static void MainO { = new SubjO; // ObsA ol = new ObsA(); ObsA o2 - new ObsAO; // // s.Register( new Del( ol.Do ) ); // s.Register( new Del( o2.Do ) ); // s.Register( new Del( ObsB.See ) ); // ( ) s.OOPSO; // Subj s } } , , . ( . : , . X . : : PS! . OOPS! , OOPS! , OOPS! object, ; . , ( ). . , Remove , : jublic void UnRegister( Del d ) dels -= d; } // 226 10. , . , . , . , , . , : Del dl = new Del( ol.Do ): // o l . D o Del d2 = new Del( o2.Do ); // o2.Do Del d3 = dl + d2; // o l . D o o2.Do d3 += d l : // o l . D o . o2.Do o l . D o d3 -= d2; // Ol.Do o l . D o , , , . , string, , , . , . : , . . , , . . (callback) , . . 10.1. , , () . , . 227 () ()- /' . 1 0 . 1 . . , W i n d o w s . 10.3. , . 1 0 . 3 . using System: namespace ConsoleApplicationl { public delegate double Fun( double x ); // class Classl { public static void Tablet Fun F, double x, double b ) { Console.WriteLineC " X Y " ); while (x <= b) { Console.WriteLineC "\| {0,8:0.000} \| {1,8:0.000} \|". x, F(x)); x += 1: } Console.WriteLineC " " ); public static double SimpleC double x ) { return 1: } static void MainO { Console.WriteLineC " Sin " ); 10. , 2 2 8 1 0 . 3 () Tablet new Fun( Math.Sin ), -2, 2 ); Console.WriteLineC " Simple " ); TableC new Fun( Simple ), 0. 3 ); } } } : Sin X Y I -2.000 I -0,909 I -1.000 I -0,841 I 0.000 I 0,000 I 1,000 I 0.841 I 2,000 j 0,909 I j j I Simple X Y I 0,000 I 1,000 I I 1.000 I 1,000 I I 2,000 I 1,000 j I 3,000 I 1.000 I Visual Studio 2005, 2.0 C # , . , , . , , . 10.4 , 10.3. 1 0 . 4 . ( 2.0) using System; namespace ConsoleAppl icationi { public delegate double FunC double x ); // class Classl { public static void TableC Fun F. double x, double b ) { Console.WriteLine( " while ( x <= b ) " ): 229 { Console.WriteLineC "\| {0.8:0.000} \| {1,8:0.000} \|", x. FCx)): x += 1; } Console.WriteLineC " " ): static void MainO { Console.WriteLineC " Sin " ): TableC Math.Sin. -2. 2 ); // 1 Console.WriteLineC " Simple " ); TableC delegate (double x ){ return 1: }, 0. 3 ); // 2 } } } , Sin, 1 . , . Simple, , . . , : , . , . 10.5. 1 0 . 5 . - using System; namespace ConsoleAppli cationl { abstract class TableFun { public abstract double F( double x ); public void TableC double x, double b ) { Console.WriteLineC " while ( x <= b ) 1 2005 # , 1992 , . & 230 10.5 () Console.WriteLineC "\| {0.8:0.000} \| {1.8:0.000} \|". , FCx)); += 1; } Console.WriteLineC " ' " ); } class SimpleFun : TableFun { public override double F( double x ) { return 1; } } class SinFun : TableFun { public override double F( double x ) { return Math.Sin(x); } } class Classl { static void MainO { \ TableFun a = new SinFunO: Console.WriteLineC " Sin " ); a.TableC -2. 2 ): a = new SimpleFunO; Console.WriteLineC " Simple " ): a.TableC 0, 3 ); } } } , , , , . , , . , . , , 231 GetlnvocationList. 10.6, 10.1. 1 0 . . using System; namespace ConsoleApplicationi { delegate void Del ( ref string s ); class Classl { public static void C001 ( ref string s ) { Console.WriteLineC " C001" ); string temp = ""; for ( int i = 0; i < s.Length; ++i ) { if ( s[i] == 'o' \|\| s[i] == 'O') temp += ' 0 ' ; else if ( s[i] == ' 1 ' ) temp += ' ; else temp += s[i]; } s = temp; } public static void Hack ( ref string s ) { Console.WriteLineC " Hack" ); string temp = ""; for ( int i = 0; i < s.Length; ++i ) if ( i / 2 * 2 == i ) temp += char.ToUpperC s[i] ); else temp += s[i]; s = temp; public static void BadHack ( ref string s ) { Console.WriteLineC " BadHack" ); throw new Exception(); static void MainO { string s = "cool hackers"; Del d = new Del C001 ) ; d += new Del( BadHack ); d += new DelC Hack ); // // // // 232 10. , 10.6 () foreach ( Del fun in d.GetlnvocationListO ) { try { fun( ref s ); // } catch ( Exception ) { Console.WriteLineC .Message ); Console.WriteLineC "Exception in method " + fun.Method.Name); } } Console.WriteLineC " - " + s ); } } } : COOl BadHack Exception of type System.Exception was thrown. Exception in method BadHack Hack - COOl hAcKeRs GetlnvocationList Method. Method Info. Method Info , , , . 12. , . , , - . - . , , . : , (sender) , , , , (receivers) , . 233 : - . : , ; (), . 1 [ ] [ ] event new, public, protected, internal, private, static, virtual, sealed, override, abstract extern, . , , (static), , . , . : public delegate void Del( object ); class // { public event Del Oops; // } - . - , . ( !), , - . , , . , , , += - = , . . , , , . 10.7 10.2, . 1 . , . 234 10. , 1 0 . 7 . using System; namespace ConsoleApplicationl { public delegate void D e l O ; // class Subj { // - // public event Del Oops; // , public void CryOopsO { Console.WriteLineC "OOPS!" ); if ( Oops != null ) OopsO: } class ObsA { public void D o O ; // - // Console.WriteLineC ", OOPS!" ); } class ObsB // - { // public static void S e e O { Console.WriteLineC " , OOPS!" ) } class Classl { static void MainO { Subj s = new Subj(); // ' ObsA ol = new ObsAC): ObsA o2 = new ObsAC); // s.Oops += new DelC ol.Do ); s.Oops += new DelC o2.Do ); s.Oops += new DelC ObsB.See ); // // // s. CryOopsO; // // 235 : , += -=. void, . . , , , , : , . . null. , CryOops nul 1 , System.Nul1ReferenceException. . N E T , . : EventHandler; : object; EventArgs . , , EventArgs, . , , System.EventHandler. On . 10.8 10.7, .NET. ! 1 0 . 8 . EventHandler using System; namespace ConsoleApplicationi { class Subj { public event EventHandler Oops; public void CryOops() { Console.WriteLineC "OOPS!" ); if ( Oops != null ) OopsC this, null ); } } class ObsA ^ & 236 10. , 10.8 () public void OnOopsC object sender. EventArgs e ) { Console.WriteLineC ", OOPS!" ); } } class ObsB { public static void OnOopsC object sender, EventArgs e ) { Console.WriteLineC " , OOPS!" ): } } class Classl { static void MainO { Subj s = new SubjC); ObsA ol = new ObsAC); ObsA o2 = new ObsAC); s.Oops += new EventHandlerC ol.OnOops ): s.Oops += new EventHandlerC o2.0nOops ): s.Oops +- new EventHandlerC ObsB.OnOops ); s. CryOopsO: } } } , # 2.0, , . 10.9. , . 1 0 . 9 . ( 2.0) using System; namespace ConsoleApplicationl { class Subj { public event EventHandler Oops; public void CryOopsO { Console.WriteLineC "OOPS!" ) 237 if ( Oops != null ) Oops( this, null ); } class ObsA { public void OnOopsC object sender, EventArgs e ) { Console.WriteLineC ", OOPS!" ); } } class ObsB { public static void OnOopsC object sender, EventArgs e ) { Console.WriteLineC " , OOPS!" ): class Classl { static void MainO { Subj s = new SubjC); ObsA ol ' new ObsAC); ObsA o2 = new ObsAC); s.Oops += ol.OnOops; s.Oops += o2.0n0ops; s.Oops += ObsB.OnOops; s.Oops += delegate ( object sender, EventArgs e ) { Console.WriteLineC " !" ); }; s.CryOopsO; } } } . N E T , , Windows.Forms, Windows- . 14. .NET . . () , 238 10. , . 1 . (thread ) . , . Main. , . . . , . , , . : , , ; , (, , , , , , ). Thread . N E T System.Threading. . 10.1. 1 0 . 1 . System.Threading Interlocked , , Monitor Mutex - , ReaderWriterLock , , Thread , , , , -, . . 239 ThreadPool Timer WaitHandle , , IOCompletionCallback , - , , ThreadStart TimerCallback , , Timer WaitCaback , ThreadPool ThreadPriority ThreadState . Thread. - , , : Thread t = new Thread ( new ThreadStartC _ ) ); , . 10.10 . 1 0 . 1 0 . using System: using System.Threading: namespace ConsoleAppl icationi { class Program { static public void HedgehogO // for ( int i = 0; i < 6: ++i ) { Console.WriteLineC i ); Thread.SleepC 1000 ); } } static void MainO { Console.WriteLineC " " + Thread.CurrentThread.GetHashCodeO ); 240 10. , 10.10 () Thread ta = new ThreadC new ThreadStart(Hedgehog) ); Console.WriteLineC " " + ta.GetHashCodeC) ); ta.StartC); for ( int i = 0; i > -6: --i ) { . Console.WriteC " " + i ); Thread.SleepC 400 ); } } } : 1 2 0 0 -1 -2 1 -3 -4 2 -5 3 4 5 Sleep, . , . , , , . . 10.2 Thread. 1 0 . 2 . Thread CurrentThread ( ) IsAlive true false , IsBackground , , Name Priority / ( ThreadPrority) ThreadState ( ThreadState) Abort ThreadAbortException. GetData, SetData () GetDomain, GetDomainID ( ), 241 GetHashCode Sleep Interrupt Join Resume Start ThreadStart Suspend . , , . 10.11. 1 0 . 1 1 . , using System; using System.Threading; namespace ConsoleApplicationi { class Classl { public void Do() { for ( int i - 0: i < 4; ++i ) { Console.Write( " " + i ); Thread.SIeep( 3 ); } } } class Program { static void MainO { Classl a = new ClassK); Thread tl = new Thread( new ThreadStartC a.Do ) ); tl.Name = "Second Console.WriteLineC " " + tl.Name ); tl.StartO; Thread t2 = new ThreadC new ThreadStartC a.Do ) ); t2.Name = "Third": Console.WriteLineC " " + t2.Name ); t2.Start(); } } } 242 10. , : Second Third 0 0 1 1 2 2 3 3 , . , lock. : lock ( ) _ , . this, typeof (). , . , Do : public void Do() { lockC this ) { for ( int i = 0; i < 4; ++i ) { Console.Write( " " + i ): Thread.SIeep( 30 ): } } } : Second Third 0 12 3 0 12 3 , , Beginlnvoke Endlnvoke. Beginlnvoke , . . Beginlnvoke , . , AsyncCal 1 back. , Endlnvoke. 'j 243 Beginlnvoke, End Invoke , . 10.11 , . Visual Studio . Factorizer Factorize, . : Numl Beginlnvoke, Num2 End Invoke. 1 0 . 1 1 . using System: using System.Threading; using System.Runtime.Remoting.Messaging; // public delegate bool AsyncDelegate ( int Num. out int ml, out int m2 ); // , public class Factorizer { public bool FactorizeC int Num. out int ml, out int m2 ) { ml = 1; m2 = Num; for ( int i = 2; i < Num; i++ ) if ( 0 == (Num % i) ) { ml = i; m2 = Num / i; break; } if (1 == ml ) return false; else return true; } } // , public class PNum { private int Number; public PNum( int number ) { Number = number; } [OneWayAttributeO] // , public void Res( IAsyncResult ar ) { int ml, m2; // AsyncResult AsyncDelegate ad = (AsyncDelegate)((AsyncResult)ar).AsyncDelegate; // Factorize ad.EndInvoke( out ml, out m2, ar ); & 244 10. , 10.11 () II Console.WriteLineC " : {0} : {1} {2}", Number, ml. m2 ); } } // public class Simple { // 1: public void NumlO { Factorizer f = new FactorizerO; AsyncDelegate ad = new AsyncDelegate ( f.Factorize ); int Num = 1000589023, tmp; // , // Factorize PNum n = new PNum( Num ); // AsyncCallback callback = new AsyncCallbackC n.Res ): // Factorize IAsyncResult ar = ad.BeginlnvokeC Num, out tmp, out tmp, callback, null ); // // - // ... } // 2: public void Num2() { Factorizer f = new FactorizerO; AsyncDelegate ad = new AsyncDelegate ( f.Factorize ): int Num = 1000589023. tmp; // , // Factorize PNum n = new PNum( Num ); // AsyncCallback callback = new AsyncCallbackC n.Res ); // Factorize IAsyncResult ar = ad.BeginlnvokeC Num, out tmp, out tmp, null, null ); // ar.AsyncWaitHandle.WaitOne( 100.00. false ); 245 if ( ar.IsCompleted ) { int ml. m2; // Factorize ad.Endlnvoket out ml, out m2. ar ); // Console.WriteLine( " : {0} : {1} {2}", Num, ml. m2 ); } } public static void MainO { Simple s = new SimpleO; s.NumlO; S.Num2(); } } : : 1000589023 : 7 142941289 : 1000589023 : 7 142941289 [OneWayAttributeO] . . N E T , , Windows. , , , , . + + , . , . 2.0, # , . . , , . 11 , . , , , , . , , , . - # - .NET. . . (stream) . , , . , ( , , ). , , . . , . . , , , . . 1 , . , ! 247 . N E T , . 11.1. System. 10. . . 1 1 . 1 . .NET , . . 11.1. 1 1 . 1 . System. BinaryReader, BinaryWriter (, , . .) BufferedStream (, ) Di rectory, Directorylnfo, File, Filelnfo : , , . Fi 1 Di rectory . Directorylnfo Filelnfo FileStream () , MemoryStream & 1 1 . 248 11.1 () StreamWriter, StreamReader ( ) StringWriter, StringReader , : (BinaryReader, BinaryWriter); (FileStream); , (StreamWriter, StreamReader). . N E T Unicode, . , , , , . , , . , . , () , , , () . , . . , , # . Convert Parse, - (. . 59). , ! , , ] ToString, . . N E T XmlTextReadf XmlTextWriter, ' X M L . X M L 15. 249 . : 1. . 2. (-). 3. . , . , , . FileAccess, System. 10. . 11.2. 1 1 . 2 . FileAccess Read ReadWrite Write FileMode (. 11.3). 1 1 . 3 . FileMode Append , , . , Create . , CreateNew . , IOException Open " OpenOrCreate , . , Truncate FileMode.Append FileAccess.Write, , . FileShare (. 11.4). 250 1 1 . 1 1 . 4 . FileShare None Read . , ReadWrite Wri te - Fi I eStream, Stream, . Stream . 11.5. 1 1 . 5 . Stream BeginRead, BeginWrite CanRead, CanSeek, , , : , / CanWrite ose (, . .) EndRead, EndWrite Flush . , Length Position Read, ReadByte ( ) Seek SetLength Wri te, WriteByte ( ) 251 FileStream . FileMode, FileAccess FileShare. . 11.211.4. 11.1 . , . 1 1 . 1 . using System; using System.10; namespace ConsoleApplication1 { class Classl { static void MainO . { FileStream f = new FileStream( "test.txt", FileMode.Create, FileAccess.ReadWrite ); f.WriteByte( 100 ); II 100 byte[] = new byte[10]; for ( byte i = 0; i < 10; ++i ) { x[1] = (byteM 10 - i ); f.WriteByte(i); // 10 0 9 f.WriteC x, 0, 5 ); // 5 byte[] = new byte[20]; f.Seek( 0, SeekOrigin.Begin ); f.Read( ,. 0. 20 ); // - // foreach byte elem in ) Consol.WriteC " " + elem ); Console.WriteLineC); f.Seek(5, SeekOrigin.Begin); // - 5- int = f.ReadByteO; // 5- Console.WriteLineC ); = f.ReadByteO; // 6- Console.WriteLineC ); Console.WriteLineC " " + f.Position ); f.CloseO; } } } 252 1 1 . 100 0 1 2 3 4 5 6 7 8 9 1 0 9 8 7 6 0 0 0 0 4 5 7 ( , Append) . Seek, : , . SeekOrigin: Begin, Current End. . , , (. . 30), : FileStream f = new FileStreamC @"D:\CJ\test.txt", FileMode.Create, FileAccess.ReadWrite ): . , , , . , , : FileNotFoundException, ; DirectoryNotFoundException, ; Argument Except ion, ; IOException, - -. . , . , : try { FileStream f = new FileStreamC @"d:\C#\test.tx", Fi1eMode.Open. FileAccess.Read ); // f.CloseO; } catchC FileNotFoundException e ) 253 { Console.WriteLineC .Message ); Console.WriteLineC " !" ); return; } catch( Exception e ) { Console.WriteLineC "Error: " + e.Message ); return; } , , , , . , , . , , Flush. Stream (, , FileStream) -: . , -. - , 1 . . - , . BeginRead. , , , , . , , , . EndRead, . . 11.2 . , , . 254 1 1 . 1 1 . 2 . using System; using System.10; using System.Threading; namespace ConsoleApplicationl { class Demo { public void UserlnputO // { string s; do { Console.WriteLineC " . Enter " ); s = Console.ReadLineO; } while (s.Length != 0 ); } public void OnCompletedReadC IAsyncResult ar ) .// 1 { int bytes = f.EndRead( ar ); Console.WriteLineC " " + bytes ); } public void AsyncReadO { f = new FileStreamC "D:\\verybigfile". FileMode.Open. FileAccess.Read, FileShare.Read. buf.Length, true ); callback = new AsyncCallbackC OnCompletedRead ): f.BeginReadC buf, 0. buf.Length, callback, null ); } FileStream f; byte[] buf = new byte[66666666]; AsyncCallback callback; } class Program { static void MainO { Demo d = new DemoO; d. AsyncReadO; d.UserlnputO; } } } 111 1/3 // 4 255 Demo. OnCompletedRead ( 1) IAsyncResult, , EndRead. , true ( 2 ) . 3 AsyncCallback, OnCompletedRead. OnCompletedRead 'BeginRead ( 4 ) , , . OnCompletedRead . verybigfile , Userlnput , OnCompletedRead. , 11.2, BeginRead EndRead, , . . StreamWriter StreamReader Unicode- , , , . TextWriter TextReader , . . 11.6 11.7 . , . 1 1 . 6 . TextWriter Close . , Fl ush System. Text.Encoding. & 256 1 1 . 11.6 {) NewLine , . (\\) Write Wri teLi 1 1 . 7 . TextReader Peek Read ReadBlock ReadLine stri ng. (nul 1) (EOF) ReadToEnd , , stri ng , : , , , . 11.3 , . . . , . ...\ConsoleApplication1\bin\Debug. 1 1 . 3 . using System; using System.10; namespace ConsoleApplicationl { class Classl { static void MainO { try { StreamWriter f = new StreamWriterC "text.txt" ); f.WriteLineC " :" ); double = 12.234; 257 int b = 29: f.WriteLineC " a = {0.6:C} b = { 1 . 2 : X } \ a. b ); f.CloseO; catch( Exception e ) { Console.WriteLineC "Error: " + e.Message ); return; } } 11.4 , , . 1 1 . 4 . using System; using System.10; namespace ConsoleApplicationi { class Classl { static void MainO { try { StreamReader f = new StreamReaderC "text.txt" ); string s = f.ReadToEndO; Console.WriteLine(s); f.CloseO: catchC FileNotFoundException e ) Console.WriteLineC e.Message ); Console.WriteLineC " !" ): return; catch( Exception e ) Console.WriteLineC "Error: return; + e.Message ); 258 1 1 . ReadToEnd. , 11.5. . 1 1 . 5 . using System; using System.10; namespace ConsoleApplicationl { class Classl { static void MainO { try { StreamReader f = new StreamReaderC "text.txt" ); string s; long i = 0: while ( ( s = f.ReadLineO ) != null ) Console.WriteLineC " { 0 } : { 1 } " . ++i, s ) ; f.CloseO; } catch( FileNotFoundException e ) { Console.WriteLineC e.Message ); Console.WriteLineC " !" ); return; } catch Exception e ) { Console.WriteLineC "Error: " + e.Message ); return; } } } } , , ] 11.6. * . : : , , * . 259 1 1 . 6 . using System; using System.10; namespace ConsoleApplicationi { class Classl { static void MainO { try { StreamReader f = new StreamReaderC "numbers.txt" ); string s: const int n = 20; i n t [ ] a = new int[n]; string[] buf; while ( s = f.ReadLineO ) != null ) { buf = s.SplitO ' ) ; long sum = 0; for ( int i = 0; i < buf.Length; ++i ) { a [ i ] = Convert.ToInt32( b u f [ i ] ) ; sum += a [ i ] ; } Console.WriteLineC "{0} ; { 1 } 0 s. sum ); } f.CloseO; } catch( FileNotFoundException e ) { Console.WriteLineC e.Message ); Console.WriteLineC " !" ): return; } catch Exception e ) { Console.WriteLineC "Error; " + e.Message ): return; } } } } : 12 4 : 7 3 44 -3 : 50 8 11 : 10 260 1 1 . , , . , . BinaryWriter , . , FileStream. . . 11.8 11.9. 1 1 . 8 . BinaryWriter BaseStream , BinaryWriter Close Flush Seek Write 1 1 . 9 . BinaryReader BaseStream , BinaryReader Close PeekChar ' Read ReadXXXX (, ReadBoolean, ReadByte, Readlnt32 . .) 11.7 . , 8888. 1 1 . 7 . using System; using System.10; namespace ConsoleApplicationl { class Classl { static void MainO I j I try { BinaryWriter tout = new BinaryWriter( new FileStreamC @"D:\C#\binary", FileMode.Create) ); double d = 0; while ( d < 4 ) { fout.WriteC d ); d += 0.33; }: fout.Seek( 16. SeekOrigin.Begin ); // fout.Write( 8888d ); fout.CloseC); } catchC Exception e ) { Console.WriteLineC "Error: " + e.Message ); return: } } } . doubl 8 . , , 11.8 , BinaryReader , . , ReadDoubl EndOfStreamException. , . 1 1 . 8 . using System; using System.10; namespace ConsoleAppl icationi { class Classl { static void MainO { try 262 1 1 . 1 1 . 8 {) '{ FileStream f = new FileStreamC @"D:\C#\binary", Fi1eMode.Open ); BinaryReader fin = new BinaryReaderC f ); long n = f.Length / 8; doublet] = new double[n]: // long i = 0: try { whileC true ) x[i++] = fin.ReadDoubleO; // } catch ( EndOfStreamException e ) {} foreachC double d in x ) Console.WriteC " " + d ); // fin.CloseO; f.CloseO; catch ( FileNotFoundException e ) Console.WriteLineC e.Message ); Console.WriteLineC " !" ); return; catch ( Exception e ) Console.WriteLineC "Error: " + e.Message ); return; } } } : 0 0,33 8888 0,99 1.32 1,65 1,98 2,31 2,64 2,97 3,3 3,63 3,96 - , . Console, System. 263 : Console. In TextReader Consol . Out Console.Error TextWriter. , , Set In SetOut ( <, > ) . , Console Read, ReadLine, Write WriteLine, In, Out Error. , . , , . System. 10 , : Directory, File, Directorylnfo Filelnfo. , , , . Directory File . Directorylnfo Filelnfo , . Directorylnfo Filelnfo FileSystemlnfo, , . 11.10. 1 1 . 1 0 . FileSystemlnfo Attributes . FileAttributes CreationTime Exists Extension Full Name LastAccessTime LastWriteTime Name . . , . , 264 1 1 . Directorylnfo , . . 11.11 1 1 . 1 1 . Directorylnfo Create, CreateSubDirectory Delete GetDirectories GetFiles Filelnfo MoveTo Parent 11.9 , , . 1 1 . 9 . Directorylnfo using System; using System.10; namespace ConsoleApplicationl { class Classl { static void DirlnfoC Directorylnfo di ) { // ===== Directory Info ): Console.WriteLineC + di.FullName ); Full Name: " Console.WriteLineC + di.Name ); Name: " Console.WriteLineC + di.Parent ) ; Parent: " Console.WriteLineC + di.CreationTime ); Creation: " Console.WriteLineC + di.Attributes ) ; Attributes: Console.WriteLineC , Root: " + di.Root ); Console.WriteLineC ); ===== Console.WriteLineC } static void MainO { Directorylnfo di1 Directorylnfo di2 new DirectoryInfo( @"c:\MyDir" ); new DirectorylnfoC @"c:\MyDir\temp" ); try { // dil.CreateO: 265 di2.Create(); // Dirlnfo(dil): DirInfo(di2); // Console.WriteLineC " { 0 } . " , dil.Name ) : dil.DeleteO; } catch Exception ) { Console.WriteLineC " " ); } } } } : ===== Directory Info ===== Full Name: c:\MyDir Name: MyDir Parent: Creation: 30.04.2006 17:14:44 Attributes: Directory Root: c:\ ===== Directory Info ===== Full Name: c:\MyDir\temp Name: temp Parent: MyDir Creation: 30.04.2006 17:14:44 Attributes: Directory Root: c:\ MyDir. , . , Delete , , : dil.DeleteC true ); // Attributes. , FileAttributes, . 11.12. 266 1 1 . 1 1 , 1 2 . FileAttributes Archive Compressed Directory Encrypted Hidden Normal , . Offline , , . , Readonly System 11.10 Filelnfo jpg d:\foto d:\temp. Exists , . 1 1 . 1 0 . using System; using System.10; namespace Consol eApplicationi { class Classl { static void MainO { try { string DestName = @"d:\tempV; Directorylnfo dest = new DirectoryInf( DestName ): dest.CreateO; // Directorylnfo dir = new DlrectorylnfoC @"d:\foto" ); if ( ! dir.Exists ) // { Console.WriteLineC " " + dir.Name + " " ); return; } FilelnfoD files = dir.GetFiles( ".jpg" ); // 267 () foreachC Filelnfo f in files ) f.CopyToC dest + f.Name ); // Console.WriteLineC " " + fi1es.Length + " jpg-" ); } catch ( Exception e ) { Console.WriteLineC "Error: " + e.Message ); } } } } Fi 1 e Di rectory , , , , . () # , . , . . , , , , . , , [Serial izable]. 12, , , . , , [NonSerialized], : [Serializable] class Demo { public int a = 1: [NonSerialized] public double y: public Monster X. Y; } : S O A P ( X M L - ) . System.Runtime.Serialization.Formatters.Binary, System.Runtime.Seri alizati on.Formatters.Soap. 268 1 1 . . BinaryFormatter, : Serialize( , ); DeserializeC ): Serialize , Deserialize . 11.11 Demo Demo.bin. , , , Visual Studic.NET. 1 1 . 1 1 . using System: using System.10; us i ng System.Runti me.Seri a1i zat i on.Formatters.Bi na ry; namespace ConsoleApplicationi { [Serializable] abstract class Spirit { public abstract void PassportO; } [Serializable] class Monster : Spirit { public Monster( int health, int ammo, string name ) { this.health = health; this.ammo - ammo; this.name = name: } override public void PassportO { Console.WriteLineC "Monster { 0 } \t health = { 1 } ammo = { 2 } " , name, health, ammo ); } string name; int health, ammo; } [Serial izable] class Demo { public int a = 1; 269 () [NonSerialized] public double b; public Monster X, Y; } class Classl { static void MainO { Demo d = new DemoO; d.X = new Monster( 100. 80. "" ); d.Y = new MonsterC 120, 50, "" ); d.a - 2; d.b = 2; d.X. PassportO; Console.WriteLineC d.a ); d. Y. PassportO; Console.WriteLineC d.b ); FileStream f = new FileStreamC "Demo.bin", FileMode.Create ); BinaryFormatter bf = new BinaryFormatterO; bf.Serialize( f, d ); // d f f.CloseO: } } . , . : Monster Monster 2 2 health = 100 ammo = 80 health = 120 ammo = 50 , : 1. System.Runtime.Serialization. Formatters.Binary. 2. [Serializable]. 3. . 4. BinaryFormatter. 5. . 6. . 11.12 . 270 1 1 . 1 1 . 1 2 . using System; using System.10; using System.Runtime.Serialization.Formatters.Binary; namespace Consol eAppl icationi { class Classl { static void MainO { FileStream f = new FileStream( "Demo.bin", FileMode.Open ); BinaryFormatter bf = new BinaryFormatterO; Demo d = (Demo) bf.DeserializeC f ); // d.X. PassportO; d.Y. Passport ; Console.WriteLineC d.a ); Console.WriteLineC d.b ); f.CloseO; } } } : Monster health = 100 ammo = 80 Monster health = 120 ammo = 50 2 0 , . , , . SOAP SoapFormatter. , ISeri al i zabl e . , , , , . , .NET. 271 , . : , . , Unicode. , . . , . . # . Convert Parse. ToString, . , , , , . . () [Serializable]. : S O A P ( XML-). 12 , , , . , . , , . . .NET dll, , , (. 12.1). 1 (. . 9 ) , . IL ( ) . 1 2 . 1 . , 273 (Intermediate Language, I L ) , , : 1 , C L R ; 2 , .NET- . [L- ILDasm.exe, ...\SDK\bin\ Visual Studio.NET. ILDasm . N E T dll ; File Open. , . , . , . . , , . , , , , , . , , . . , , , . , , , , , . , . , (). , . , . , , , , . .NET , . , ( ), ( ). . 274 12. , , , , ; , (, ); (strong name) , ; , ; , ( . N E T . [27]). . . : , : (major version); (minor version); (build number); (revision number). , . , 3.1.0.0, 3.4.0.0, , , . , . , , (patch). , [AssemblyVersion], . 285. , . . , , , , , . . . Visual Studio.NET . , . , , . 275 , (Global Assembly Cache, G A C ) . (strong name), . "' Visual Studio.NET Class Library ( ). 8 . , dll. MonsterLib (. 12.2). . 1 2 . 2 . 12.1. 8 public , . 1 2 . 1 . namespace MonsterLib { using System; public abstract class Spirit { public abstract void PassportO; & 276 12 . , , , 12.1 () public class Monster : Spirit { public Monster'O this.health = 100; this.ammo = 100: this.name = "Noname"; public MonsterC string name ) : thisO { this.name = name; } public Monster( int health, int ammo, string name ) { this.health = health; this.ammo = ammo; this.name = name; } public int Health { get { return health; } set { if ( value > 0 ) health = value; else health -= 0; } } public int Ammo { get { return ammo; } set { if ( value > 0 ) ammo = value; else ammo = 0; } } public string Name { 277 pet return inSTfe , - } } override public void PassportO { Console.WriteLineC "Monster {0} \t health = {1} ammo = { 2 } " , name, health, ammo ); } string name; int health, ammo; } public class Daemon : Monster { public Daemon() brain = 1 ; public DaemonC string name, int brain ) : baseC name ) this.brain = brain; public DaemonC int health, int ammo, string name, int brain ) : baseC health, ammo, name ) this.brain = brain; override public void PassportO Console.WriteLineC "Daemon {0} \t health = {1} ammo = {2} brain = { 3 } " . Name, Health, Ammo, brain ); public void ThinkO Console.WriteC Name + " is" ); for ( int i = 0; i < brain; ++i ) Console.WriteC " thinking" ); Console.WriteLineC " . . . " ) ; int brain; 278 12. , , , , MonsterLib.dll ...\bin\ Debug ...\obj\Debug. MonsterLib.dll ILDasm.exe, (. 12.3). . 1 2 . 3 . ILDasm.exe , . , , (. . 178), MonsterLib.dll. , ( , ) Project Add Reference ( ). , , Browse. , . 12.2. 279 1 2 . 2 . using System; namespace ConsoleApplicationi { using MonsterLib: class Classl { static void MainO { const int n = 3; MonsterC] stado = new Monster[n]: stado[0] = new MonsterC "Monia" ); stadoll] = new MonsterC "Monk" ); stado[2] = new Daemon ( "Dimon", 3 ); foreach ( Monster elem in stado ) elem. PassportO; for ( int i = 0; i < n; ++i ) stado[i].Ammo = 0; Console.writeLineO; foreach ( Monster elem in stado ) elem.PassportO; } } 8.3. ...\bin\Debug , MonsterLib.dll, , . , , . , . .NET , , . , , , VB.NET. . , , , . . . System. System.Reflection. 280 12. , , , , . System.Reflection , , , . . 12.1. 1 2 . 1 . IsAbstract, IsArray, IsNestedPublic, IsClass, IsNestedPrivate, IsCOMObject, IsEnum, Islnterface, IsPrimitive, IsSealed, IsValueType , (, , , . .). GetConstructors, GetEvents, GetFields, Get Interfaces, GetMethods, GetMembers, GetNestedTypes, GetProperties , (, , . .). , , GetFields Fieldlnfo, GetMethods Method Info. ( S ), (, GetMethod GetMethods) FindMembers Member Info GetType InvokeMember . , new , : 1. object GetType, , . , : Monster X = new MonsterC); Type t = X.GetTypeO; 2. Type GetType , (), : Type t = Type.GetTypeC "Monster" ); 3. typeof , , : Type t = typeof Monster ); 281 . . 12.1, (, Memberlnfo). System. Ref lection. . 12.2. 1 2 . 2 . System.Reflection Assembly AssemblyName (, , , . .) EventInfo Fieldlnfo , Eventlnfo, Fieldlnfo, Methodlnfo Propertylnfo Memberlnfo Methodlnfo Module Parameterlnfo Propertylnfo 12.3 12.1. 1 2 . 3 . using System; using System.Reflection; namespace ConsoleApplicationl { using MonsterLib; class Classl { static void InfoC Type t ) { Console.WriteLineC " = = = = " + t.FullName ) ; if ( t.IsAbstract ) Console.WriteLineC "" ); if ( t.IsClass ) Console.WriteLineC "" ); if ( t.IsEnum ) Console.WriteLineC "" ); Console.WriteLineC " " + t.BaseType ); MethodInfo[] met = t.GetMethodsO; foreach ( Methodlnfo m in met ) Console.WriteLineC m ); Console.WriteLineC): 282 12. , , , 12.3 () PropertyInfo[] prs = t.GetPropertiesO; , foreach ( PropertyInfo p in prs ) Console.WriteLineC p ); Console.WriteLineC); } static void MainO { Type t = typeof( Spirit ); InfoC t ); t = typeof( Monster ): InfoC t ); t = typeof( Daemon ); InfoC t ); } } : ======= MonsterLib.Spirit System.Object Void PassportO Int32 GetHashCodeO Boolean Equals(System.Object) System.String ToStringO System.Type Get Type ======= MonsterLib.Monster MonsterLib.Spirit Void PassportO Int32 GetHashCodeO Boolean Equals(System.Object) System.String ToStringO Int32 get_Health() Void set_Health(Int32) Int32 get_Ammo() Void set_Ammo(Int32) System.String get_Name() System.Type GetType() Int32 Health Int32 Ammo System.String Name 283 ======= MonsterLib.Daemon MonsterLib.Monster Void PassportO Int32 GetHashCodeO Boolean Equals(System.Object) System.String ToStringO Void Think Int32 getJealthO Void set_Health(Int32) Int32 get_Ammo() Void set_Ammo(Int32) System.String get_Name() System.Type GetTypeO Int32 Health Int32 Ammo System.String Name , , . , . , . (, , . .). . System. Attribute. . .NET , . , , , , . () , , . , . . , , . : [Serializable] class Monster 284 12. , , , [NonSerialized] string name; int health, ammo; } [Serializable], , , Monster. name [NonSerialized], , . 10. , , . , . , , . 12.3. 1 2 . 3 . , assembly field event method pa ram property return type , , ABC: [ABC] public void Do() { ... } . , , , : [return:ABC] public void Do() { ... } . . = , . , CLSCompliant true. , 285 , using, : using System; [assembly:CLSCompliant(true)] namespace ConsoleApplicationl { ... [CLSCompliant] , C L S (Common Language Specification) . , , , , () . , , , , . [STAThread], , , . , , . , ["Conditional!, . Assemblylnfo.cs, . [AssemblyVersion], : [assembly: AssemblyVersion("1.0.0.0")] . [27]. , . : , ; , . , , , . , ( ), . , , , . 286 12. , , , , : namespace State { namespace City { } } : namespace State.City { } , , .NET. , - : 1. . , System.Runtime.Serialization.Formatters.Binary BinaryFormatter. : System.Runtime.Serialization.Formatters.Binary.BinaryFormatter bf = new System.Runtime.Seri ali zati on.Formatters.Bi nary.Bi naryFormatter(); 2. using, ; . , using System.Runtime.Serialization.Formatters.Binary; BinaryFormatter bf = new BinaryFormatterO; using , ; . 3. . using: using BinF = System.Runtime.Seri ali zati on.Formatters.Bi nary.Bi naryFormatter; BinF bf .= new BinFO; , , , . 287 # 2.0 : : , : SI0 = System.10: ig MIO = MyLibrary.IO; ss Program // // static void MainO { SI0::Stream s = new MIO: .-Empty St ream ; // , ( . :: )bal. , , , . : , , ; . , , . , ) , , . + + , . # , - + + , , . , , . , . / //. . 12.4. ' . ^ , . , , . 288 12. , , , 1 2 . 4 . fdefine, fundef (, #def i ne DEBUG) (fundef DEBUG) , . . #if,#elif, felse, fendif . , , , (. ) #line , , (, #1 i 200 "ku_ku.txt"). , 1 , terror, fwarning , . f error (, ferror ). fwarning #region, fendregion , . fpragma # 2.0. (#pragma warning disable) (#pragma warning restore ) #if [ #e1if ...] [ #elif ...] [ #else ...] #endif # e l i f . , . , ==, ! =, !, && 11. . true, #def i ne. 1 ... , . . . . 289 12.4. 1 2 . 4 . // #define VAR1 // #define VAR2 using System; namespace ConsoleApplicationi { class Classl { #if VAR1 static void F ( ) { Console.WriteLine( " 1" ): } #elif VAR2 static void F(){ Console.WriteLineC " 2" ); } #el se static void F ( ) { Console.WriteLineC " " ); } #endif static void MainO { FO: } } , , F. #def i . Conditional . , . 12.5. , System.Diagnostics. 1 2 . 5 . Conditional // #define VAR1 fdefine VAR2 using System; using System.Diagnostics: namespace ConsoleApplicationi { class Classl { [Conditional ("VAR1")] static void A ( ) { Console.WriteLineC " A" ); } [Conditional ("VAR2")] & 290 12. , , , 12,5 () static void ( ) { Console.WriteLineC " " ); } static void MainO { : BO; } Main , , VAR1 . , : , , , . , [20], [21], [26], [27], [30]. 13 , - 3 , , . N E T . , , # 2.0, - 1 [generics), (partial types) (nullable types) . , . , . : , , , , , -, . . . , , . . , , . , . . 1 , 2.0, . , 9, 10. 292 13. , - , . . , . , , . . , , , , , : , , , . . : , , , . , . , , . . . , L I F O (Last In First Out, ). , . , . , , . , , . . . , F I F O (First In First Out, ). , -. , , , , -. , , . . . 293 . 13.1 ( ). , , . , . , . . 1 3 . 1 . , , , . . , . , , . , : ,, . , , . . -, , , , . 294 13. , - , , (. 13.1). . (-), -. 1 3 . 1 . - boy girl dog ' -, . - , , ( hash , ). - , , . - -, . -:, - , - (). , , . - , . , - -. - . , . , . , , . . , , , . , . - , 295 System.Collections . , . , , , . . , . , . , , , . . . N E T , . , , System.Collections, System. Col 1 ecti ons. Speci al i zed System. Col 1 ecti ons. Generi ( 2.0). . System. Array, , System.Array (. 6. 133). System.Collections System.Collections , . . 13.2 , . N E T (. . 198). 1 3 . 2 . System.Collections ICol 1 ecti on I Compare (, ) IDictionary IDictionaryEnumerator , IDictionary 296 13. , - 13.2 () IEnumerable IEnumerator IEnumerator foreach IHashCodeProvider I Li st . 13.3 , 1 System. Collections . 1 3 . 3 . System.Collections ArrayList IList, ICollection, IEnumerable, ICloneabl BitArray I Col lection, IEnumerable, ICloneable Hashtable Queue IDictionary, ICollection, IEnumerable, ICloneable ICollection, ICloneable, IEnumerable SortedList , . IDictionary, ICollection, IEnumerable, ICloneable Stack ICollection, IEnumerable System.Collections.Specialized , StringCollection - StringDictionary. ArrayLi st. ArrayList , , , . ArrayList . 1 2 13.2 13.3 [27] . , Hashtabl System.Object.GetHashCode. 297 System.Collections ArrayLi st 16 object. , Capacity, : ArrayList arrl = new ArrayList(); // 16 ArrayLi st arr2 = new ArrayList(1000); // 1000 ArrayList = new ArrayList(); arr3.Capacity = 1000; // ArrayLi st . 13.4. 1 3 . 4 . ArrayList Capacity ( , ) Count Item Add AddRange BinarySearch Clear Clone CopyTo GetRange ArrayLi st IndexOf ( -1, ) Insert ( ) InsertRange LastIndexOf Remove RemoveAt RemoveRange Reverse SetRange Sort TrimToSize (. . 205). 298 13. , - ArrayLi st Array, . # object, . , , object, -. , , . , , ArrayLi st. , , , . ArrayLi st: arrl.AddC 123 ) ; arrl.Add( - 2 ) : arrl.AddC "" ); , , : int = (int) arrl[0] int b = (int) arrl[l] string s = (string) a r r l [ 2 ] , , InvalidCastException. : ArrayLi st , " , , ArrayLi st. ~ 13.1, Monster . (. . 178), ~ , Stado . 1 3 . 1 . using System; using System.Col lections; namespace ConsoleApplicationl { class Monster { . . . } class Daemon : Monster { . . . } class Stado : IEnumerable { 299 private ArrayList l i s t ; public StadoO { l i s t = new ArrayListO; } public void Add( Monster m ) { list.Add( m ) ; } public void RemoveAtC int i ) { list.RemoveAtt i ); } public void ClearO { list.ClearO; } public IEnumerator GetEnumeratorO { return list.GetEnumeratorO; } } class Classl { static void MainO { Stado stado = new StadoO; stado.Add( new Monster( "Monia" ) , ) ; stado.Add( new MonsterC "Monk" ) ); stado.Add( new Daemon ( "Dimon", 3 ) ); stado. RemoveAU 1 ); foreach ( Monster x in stado ) x. PassportO; } } } : Monster Monia Daemon Dimon health = 100 ammo = 100 health = 100 ammo = 100 brain = 3 , -, . , . # , 2.0, - (generics), . . - , . . - (generics) , . , , . generics -: , , , , , - . - , , . 300 13. , - . N E T , , , , , . . , System.Collections.Generic, System.Collections, System.CoIlections (. . 295). . 13.5 . N E T (, , , ). 1 3 . 5 . .NET 2.0 - ( 2.0) Comparer<T> Comparer Dictionary<K,T> HashTable LinkedList<T> List<T> ArrayList Queue<T> Queue SortedDi cti ona ry<K,T> SortedList Stack<T> Stack , . N E T 1.0 1.1, , , object: , , , ; , . - , . . ArrayLi st List<T> Monster Daemon, 1 5 8, . 1 3 . 2 . List<T> using System; using System.Collections.Generic; using System.Text: (. . 275) 301 namespace ConsoleAppl icationi { using MonsterLib; class Program { static void MainO { List<Monster> stado = new List<Monster>(); stado.Add( new MonsterC "Monia" ) ); stado.Add( new MonsterC "Monk" ) ); stado.AddC new Daemon ( "Dimon", 3 ) ); foreach ( Monster x in stado ) x.PassportO; List<int> lint = new List<int>(); lint.AddC 5 ); lint.AddC 1 ); lint.AddC 3 ); lint.SortO; int a = l i n t [ 2 ] ; Console.WriteLineC a ); foreach ( int x in lint ) Console.WriteC x + " " ); } } } : Monster Monia Monster Monk Daemon Dimon 5 1 3 5 health = 100 ammo = 100 health = 100 ammo = 100 health = 100 ammo = 100 brain = 3 13.2 . (stado) , MonsterL.ib.clll, . , Monster, , . , , , , , , . , , , . 1 i nt , , , (. 13.1). - , , 302 13. , - . , -, . , . . 13.3 . , . Dictionary<T,K>. : , . , , , , . 1 3 . 3 . using System: using System.Collections.Generic; using System.Text; using System.10; namespace ConsoleApplicationl class Program { static void MainO StreamReader f = new StreamReader( @"d:\C#\text.txt" ); // string s = f.ReadToEndO; // // 3 char[] separators = { ' ', ' ! ' }; List<string> words = new List<string>( s.Split(separators) ); / / 4 Dictionary<string, int> map = new Dictionary<string. i n t > ( ) ; / / 5 foreach ( string w in words ) // if ( map.ContainsKeyC w ) ) map[w]++; else map[w] = 1; foreach ( string w in map.Keys ) Console.WriteLineC " { 0 } \ t { l } " , w. map[w] ) ; // 7 303 text. txt . , . , ! : 1 3 1 2 4 1 2 1 2 1 2 1 1 1 . 1 , 32 767 , 2 . , . , , , FileNotFoundException (. 11). 3 , Spl it, , . words List<string>. . 5 , 6 words. , , , . , . 7 ( Keys, ) . Split : , , . , 15. 304 13. , - , . , , . , System.Collections.Generic , . 13.6. 1 3 . 6 . .NET 2.0 ( 2.0) ICollection<T> ICollection IComparable<T> IComparable IDi ct i ona ry<K. IDictionary IEnumerable<T> I Enumerable IEnumerator<T> I Enumerator IList<T> IList C# - , , , (generic) . - , # . , , , , : public class Stack<T> { [ ] items: int count; public void PushC T item ) { . . . } public T PopO { ... } // // } , int: Stack<int> stack - new Stack<int>(); stack.PushC 3 ); int x = stack.PopO; Stack<int> (constructed type). 305 . Stack , doubl , . , , . - . - . (constraints), , , , , , , . where, : public class Stack<T> where T : struct { ... struct , . class. , , , , , . , , . , - - , , , . . , - , -. new(), : public class EntityTabl<,> where : IComparable<K>, IPersistable where E: Entity. new() { public void Add( key. E entity ) { if ( key.CompareToC x ) < 0 ) { ... } } } EntityTable , , Entity . 306 13. , - , , , . - default. null, 0. , - . . , . , , . , , , . 13.4 . , , , , , . ., - 1 . . 1 3 . 4 . using System; using System.Collecti ons.Generi ; using System.Text; namespace ConsoleApplicationl { class Program static void Sort<T> ( ref T [ ] a ) where T : IComparable<T> // 1 // 2 { T buf; int n = a.Length; for ( int i = 0; i < n - 1: ++i ) { int im = i; for ( int j = i + 1; j < n; ++j ) if ( a[j].CompareTo(a[im]) < 0 ) im = j; buf = a [ i ] ; a [ i ] = a[im]; a[im] = buf; } } static void MainO II 307 i n t : ] = { 1, 6. 4, 2, 7. 5, 3 } ; Sort<int>( ref ); // 4 foreach ( int elem in aa )) Console. Console.WriteLineC elem ); double[] b = { 1.1, 5 2, 5.21, 2, 7, 6, 3 } : SortC ref b ); foreach ( double elem in b ) Console.WriteLineC elem ); // 5 string[] s = { "qwe", qwer", "df", "asd" }; SortC ref s ); foreach ( string elem in s ) Console.WriteLineC elem ); // 6 , H a i ( 2 ) , - CompareTo, 3. ( Main) Sort : - ( 4) ( 5 6). , . - , . , : , ; , . , - , . , , . -, , , , . , . 308 13. , - , (partial types). , , , , . , , . partial. , , : public partial class { } public partial class A { } , . , , . partial class, struct interface . . parti al , , . . abstract sealed, . - , . , : , . . , , , . 309 , , . . , - , . , , - 1 . , . , , . , , , . # 2.0 , (nullable). , ( Value) , , ( HasValue). , HasValue true. null, HasValue false, Value . , ?, : int? = 123; int? = null; if ( .HasValue ) Console.WriteLineC x ); // x x.Value if ( y.HasValue ) Console.WriteLineC ); , , : int i = 123: int? = i: double? = x; int? z = (int?) y; int j = ( i n t ) z; //int - - > int? // int? - - > double? // double? - - > int? // int? - - > int . == != true, null. , nul 1 . <, >, <= >= false, nul 1. nul 1, nul 1, : int? = null; int? = + 1; // = null 310 13. , - ?? (null coalescing operator). , , null, . , null, : int? = null: int = ?? 0; = 1: = ?? 0: // = // = 1 X M L . , , . , , . , . .NET . , 2.0 , , 2.0, . , , - . , C L R . - , . . , , . 14 Windows # , .NET, . Windows- , . Windows . . : , . , . Windows . . , : . . . . , . . , . . 312 14. Windows , , A P I (Application Program Interface ). API- (Dynamic Link Library, D L L ) , j . , , . Windows , , . , . , . . \| \ 4 . Windows , , Windows 95/98 , Windows NT/2000. . , O L E (Object Linking and Embedding ) . . 32- W i n d o w s 16- Windows-, , M S - D O S . DOS-, , M S - D O S . ( ). Windows . , , . ( , , ) . , , . 14.1. 1 32- , , 32 . 313 . 1 4 . 1 . , W i n d o w s , (, , ). , , . , , , . , . , , (. 14.2). , Windows- , , . . 1 4 . 2 . Windows- Visual Studio.NET Windows- , , , . . . 314 14. Windows Windows- (File New Project), Windows Application (. 14.3). , , Windows-. , , Form1.cs[Design], . ( ) , , , , . . . 1 4 . 3 . , . , Solution Explorer (View Solution Explorer) Forml .cs View Code. , , , 14.1. , , , . 1 4 . 1 . Windows- using using using using using using System; System.Drawing: System.Col lections; System.ComponentModel; System.Windows.Forms; System.Data; 315 1 Windows- nespace WindowsApplicationl public class Forml : System.Windows.Forms.Form { private System.ComponentModel.Container components = null; public Forml() { InitializeComponent (); } protected override void Dispose( bool disposing ) { i f ( disposing ) { if (components != null) { components.Di spose(); } } base.Dispose( disposing ); } #region Windows Form Designer generated code private void InitializeComponent { this.components = new System.ComponentModel.ContainerO; this.Size = new System.Drawing.SizeOOO.300): this.Text = "Forml"; } #endregion static void MainO { Application. Run (new FormlO); } } .NET. , , : jsing System; jsing System.Windows.Forms; . System . System.Windows.Forms , Windows-. 316 14. Windows . 14.1, . 14.4. 1 4 . 1 . Windows.Forms Application Windows-. Windows-, . . ButtonBase, Button, CheckBox, ComboBox, OataGrid, GroupBox, ListBox, LinkLabel, PictureBox , (): , , , , , , , , Form Windows- ColorDialog, FileDialog, FontDialog, PrintPreviewDialog , , , Menu, MainMenu, Menultem, ContextMenu Clipboard, Help, Timer, Screen, ToolTip, Cursors : , , , , , StatusBar, Splitter, ToolBar, ScrollBar , : , , . . Wi ndows. Forms . 14.1. Forml, Form. , . Forml components , . InitializeComponent, ( fregion fendregion). , . , ; (Properties), : this.BackColor = System.Drawing.SystemColors.AppWorkspace: Di spose ] . , Mai , ^ Run Application. Run , , Windows- 317 . 1 4 . 4 . Windows.Forms , , D e b u g S t a r t F5. , Windows-: , . Windows- : 1. , . 2. . ( ) . , 318 14. Windows View Properties Window. , . , (. 14.5). . 1 4 . 5 . Toolbox ( , View Toolbox), . . , . ( ). , . +, , . . , Windows Forms, . 14.6. (. . 325). , , , . ., , , . 319 Windows- . 1 4 . 6 . . . , Events , ( Events , . 14.5). , . : Acti vated ; Click, Doubleclick ; Closed ; Load ; KeyDown, KeyUp ; Keypress , ASCII-; MouseDown, MouseUp ; MouseMove ; Paint . 14.2 , , . 14.5, Click, KeyPress. . 1 4 . 2 . using System; using System.Drawing: using System.Collections: & 320 14. Windows 14.2 {) using System.ComponentModel; using System.Windows.Forms; using System.Data; namespace WindowsApplicationi { public class Forml : System.Windows.Forms.Form // // // 1 2 // // // 3 4 5 { private System.Windows.Forms.TextBox textBoxl; private System.Windows.Forms.Button buttonl; private System.ComponentModel.Container components = null; public Forml { 1 i t i a1izeComponent(): protected override void DisposeC bool disposing ) { . . . } #region Windows Form Designer generated code private void InitializeComponentO { this.textBoxl = new System.Windows.Forms.TextBoxO: this.buttonl = new System.Windows.Forms.ButtonO; this.SuspendLayoutO; // // textBoxl // this.textBoxl.Location = new System.Drawing.Point(24, 16); this.textBoxl.Name = "textBoxl": this.textBoxl.Size = new System.Drawing.Size(240. 20); this.textBoxl.Tablndex = 0; this.textBoxl.Text = "textBoxl"; this.textBoxl.KeyPress += new // 6 System.Wi ndows.Forms.KeyPressEventHandler(thi s.textBoxl_KeyPress); // // buttonl // this.buttonl.Location = new System.Drawing.Point(192. 80); this.buttonl.Name = "buttonl"; this.buttonl.Tablndex = 1 ; this.buttonl.Text = "buttonl"; this.buttonl.Click += new 111 System.EventHandler(this.buttonl_Cl ick); // // Forml // this.AutoScaleBaseSize = new System.Drawing.Size(5. 13); this.ClientSize = new System.Drawing.Size(292, 126): this.Controls.Add(this.buttonl); // 321 Windows- thi s.Controls.Add(thi s.textBoxl) this.Name = "Forml"; this.Text = "Forml"; this.ResumeLayout(false); // 9 // 10 } lendregion static void MainO Application.Run(new Forml 0 ) ; private void buttonl_Click(object sender. System.EventArgs e) // 11 private void textBoxl_KeyPress(object sender. System.Windows.Forms.KeyPressEventArgs e) // 12 } . 1 2 : TextBox Button. System.Windows .Forms, using, : public class Forml : Form // { private TextBox textBoxl; // 1 private Button buttonl; 111 private Container components = null; , , . , Name . Class View (View Class View), Name . , buttonl , . , . InitializeComponent. 3 4 , , , , . . 322 14. Windows 6 7. . , 10 (. . 232) . , buttonl, ! ick, buttonl_Click, Forml. , ( ) . . , : - , . , Events . , Controls ( 8 9). , . (Add Remove). , : 1. . 2. , . 3. . 5 10 , . 5 , 10 . ( 11 12) , . , : MessageBox.Show(" buttonl"); MessageBox.Show(" " + e.KeyChar); // 11 // 12 Show MessageBox, System.Windows.Forms. , , . , , Control. 323 Control Control Control , , , , , . Control . , , , . , Parent, , 1 , , . Control . 14.2, . 14.3. 1 4 . 2 . Control Anchor , . , BackColor, Backgroundlmage, Font, ForeColor, Cursor : , , , , Bottom, Right . Size Top, Left . Location Bounds Rectangle (), CIientRectangle Rectangle, ContextMenu Dock Location , , Pol nt. X Y Height, Width Size Size. Height Width , . , , . & 324 14. Windows 14.2 {) Created, Disposed, Enabled, Focused, Visible bool, : , , , , Handle ( , ) ModifierKeys , (Shift, Control, Alt). Keys MouseButtons , . MouseButtons Opacity . 0 () 1 () Parent , ( , -) Region Region, Tablndex, TabStop Tab , 1 4 . 3 . Control Focus GetStyle, SetStyle . Control Styles (. ) Hide, Show Visible (Hide , Show ) Invalidate QnXXXX . , , , - (OnMouseMove, OnKeyDown, OnResize, OnPaint . .), Refresh SetBounds, SetLocation, SetClientArea , , . 325 Control Styles , . , ResizeRedraw. , . , - , . . 14.4 , Control. 1 4 . 4 . Control Click, Doubleclick, MouseEnter, MouseLeave, MouseDown, MouseUp, MouseMove, MouseWheel KeyPress, KeyUp, KeyDown BackColorChanged, ContextMenuChanged, FontChanged, Move, Paint, Resize GotFocus, Leave, LostFocus , , . : . , Events . , . , OnXXXX (OnMouseMove, OnKeyDown, OnResize, OnPaint . .), , - . [27]. , , ToolBox (View ToolBox). . . 326 14. Windows Label . Text. ( Font), (BackColor), (ForeColor) (TextAlign) . (AutoSize = True). (Image) ( BackColor Col or. Transparent). , , . (. . 327). Button , , (Click). , , . . Enter , , . , Text, , . AcceptButton , , Enter Click, . . , Cancel Button , , Esc Click . . , (, ). . , , . , ( Yes), , , Cancel. 1 4 . 5 . DialogResult None Ignore Ignore OK Yes Yes Cancel Cancel No No Retry Retry Abort Abort 327 , , DialogResult. DialogResult, System. Wi ndows. Forms. . 14.5. . TextBox TextBox , Text. ( 32 ), , () , ( PasswordChar). Multiline, Scrol 1 Bars Wordwrap. Readonly. (Clear), (Select), (), (Paste) ., , KeyPress KeyOown. , Label, Button TextBox. . 14.7. . 1 4 . 7 . Enter, , . , !. . , (. 14.8). , . . , . Text . 14.6 ( ; ). 328 14. Windows . 1 4 . 8 . 1 4 . 6 . Text Text Forml label 1 label2, label3, textBoxl buttonl . , , . , ( Load) ( CI i ck). Enter ( KeyPress textBoxl). Events (. . 14.5). i . max . rnd 2 . 14.3. 1 4 . 3 . using System; using System.Drawing; using System.Collections; using System.ComponentModel; using System.Windows.Forms; using System.Data; 1 2 , . Random (. . 148). 329 namespace WindowsApplicationl { public class Forml : Form { private Label labell: private TextBox textBoxl; private Button buttonl; private Label label2; private Label label3; private Container components = null; public Forml { ... } protected override void Dispose( bool disposing ) { . . . } Windows Form Designer generated code { ... } const int max - 10; Random rnd; int i, k; // // II static void MainO { ... } private void Forml_Load(object sender, EventArgs e) { rnd = new RandomO; i = rnd.Next(max); // 0 max } private void textBoxl_KeyPress(object sender, KeyPressEventArgs e) { int n; if ( e.KeyChar != (char)13 ) return; // Enter, try { // n = Convert.Int32(textBoxl.Text); } catch { n = -1; // , } if ( n != i ) // = = = = = = = = = = { label 2.Left += 5; label2.Text = "He !"; textBoxl. ClearO; k++; // buttonl.Visible = false; } 330 14. Windows 14.3 () else { 1abel2.Left = 32: // 1abel2.Text = " "; double koef = 1.0 * k / max: label3.Text = koef.ToStringO; buttonl.Visible = true; } private void buttonl_Click(object sender. System.EventArgs e) { i = rnd.Next(max) k = 0; textBoxl.CI ear() textBoxl.Focus label 2.Text = "" 1abel3.Text = "" // 0 max // // // // MainMenu ContextMenu MainMenu , : Toolbox. , . Menultem, Text. , Enter . , , Name , . ( Enabled), (Visible), (Checked). . Exit, . Close , Exit Appl i cati on, : private void Exit_Click(object sender, EventArgs e) { // - Exit CloseO: // : // Application.ExitO; 331 . , . , , , . ContextMenu ( , ). ContextMenu . CheckBox - - . , , Checked, true false. , . , , , , . CheckState, Checked, Unchecked Intermediate. , ThreeState, . , . Appearance : (Normal), (Button), . , , . . RadioButton , . ( Checked), . , , . , . , . , Group Panel. Appearance : (Normal), (Button), 1 . . 1 AutoCheck false . 332 14. Windows GroupBox GroupBox , , , . 14.4 , , . .' [-10; 10], [-100; 100]. / . . 14.9. ( Name) , . GroupBox. Checked radioButtonl, , maxtextBox numPosittextBox Readonly ( ). " i e !!!\| { < tffltlOl 00; j \| J . 1 4 . 9 . . Format, , Shift Ctrl. , , Format Align Lefts. , CI ick. 1 4 . 4 . using System; using System.Drawing; . using System.Collections; using System.ComponentModel; 333 using System.Windows.Forms; using System.Data; namespace WindowsApplicationl { public class Forml : Form { private Label labell; private GroupBox groupBoxl; private RadioButton radioButtonl; private RadioButton radioButton2; private TextBox numtextBox; private CheckBox numPositcheckBox; private Button createbutton; private Button calcbutton; private TextBox maxtextBox; private TextBox numPosittextBox; private TextBox arraytextBox; private CheckBox maxcheckBox; private Container components = nul1; public FormlO { . . . } protected override void Dispose( bool disposing ) { Windows Form Designer generated code { . . . } // i n t [ ] arr; static void MainO { . . . } private void createbutton_Click(object sender, EventArgs e) { Random rnd = new RandomO; int a = -10. b = 10; if ( radioButton2.Checked ) a = // -100; b = 100; // int n = 0; try { n = int.Parse(numtextBox.Text); // } catch { MessageBox.Show(" !"); numtextBox.CI e a r ( ) ; numtextBox.Focus(); & 334 14. Windows 14.4 () arraytextBox.ClearO: maxtextBox.ClearO; numPosittextBox.ClearO; // if ( n < 0 ) n = -n; // = new i n t [ n ] ; // for ( int i = 0; i < n; ++i ) { a r r [ i ] = rnd.Next(a. b ) ; // arraytextBox.Text += " " + arr[i.]; // } } private void calcbutton_Click(object sender, EventArgs e) { int max = arr[0]; int numPosit = 0; for ( int i = 0; i < arr.Length; ++i ) { if ( a r r [ i ] > max ) max = a r r [ i ] ; // if ( a r r [ i ] > 0 ) ++numPosit; // } if ( maxcheckBox.Checked ) maxtextBox. Text = max.ToStringO; else maxtextBox.Text = " " ; if ( numPositcheckBox.Checked ) numPosittextBox.Text = numPosit.ToStringO; else numPosittextBox.Text = " " ; ListBox , ( Sel ecti onMode One) ( Sel ecti onMode Multi Simple Multi Extended). SelectionMode MultiSimple, . Multi Extended Shift, Ctrl, Windows. , SelectionMode, None. , . ( MultiColumn) (Sorted = True). . Items, . Items 335 wjd, AddRange Insert. Remove RemoveAt, / . Selectedltems SelectedIndices, . 14.5 , Li stBox , , . . 14.10. " Forml \| _\|[ SystemObject SystemMashalByRefObject System. ComporrertModeLComponent System. Windows.Forms.Conttol System. Windows.Forms.ScrollableControl System.Windows.Foims.ContainerContiol System. Windows.Forms.Form . 1 4 . 1 0 . 1 4 . 5 . using System; using System.10; using System.Drawing; using System.Col lections; using System.ComponentModel; using System.Windows.Forms; using System.Data; using System.Collections.Specialized; namespace WindowsApplicationi { public class Forml : Form { private ListBox listBoxl; private Button buttonl; private Container components = null; public FormlO { . . . } protected override void DisposeC bool disposing ) { . . . } Windows Form Designer generated code 1 { . . . } 3 ' & 336 14. Windows 14.5 () static void MainO { ... } private void Forml_Load(object sender. EventArgs e) { try { StreamReader f = new StreamReaderC "input.txt" ); string buf; while ( ( buf = f.ReadLineO ) != null ) // listBoxl.Items.Add(buf); // } catch ( FileNotFoundException exc ) { MessageBox.Show( exc.Message ); return; } } private void buttonl_Click(object sender. EventArgs e) { StreamWriter f = new StreamWriter( "output.txt" ); foreach ( string item in listBoxl.Selectedltems ) f.WriteLine(item); // f.CloseO; } } } , . . N E T . , , 1 , . . , , F1 ...overview (). Remarks Example, ...Members ( ), , . , Form Application, . 1 , , , < 337 ; Form . Form : Object-^MarshalByRefObject-Component->Control- HableControl>ContainerControl. ia . . , 1 . , , - . , , Cancel, . MessageBox, Windows- (. . 322). , , , . , i - . . ( Main). . i (Multiple Document nterface, M D I ) , . . , , . . , , . MarshalByRefObject , , . Component , . , Di spose, , . , , . 1 - - . 338 14. Windows Control, , . ScrollableControl , . AutoScrol 1 AutoScrol IMinSize , . ContainerControl . , , . ., . , , TabStop Tablndex. Form Form , . , , . 14.7-14.9. 1 4 . 7 . Form AcceptButton , Enter ActiveMDIChild, IsMDIChild, IsMDIContainer ( M D I ) AutoScale , , , , , FormBorderStyle ( FormBorderStyle) Cancel Button , Esc Control Box , , Menu, MergedMenu MaximizeBox, MinimizedBox 339 nTaskbar , Windows Position , ( FormStartPosition). , CenterScreen wState ( FormWi ndowState) 1 4 . 8 . Form vate :erToScreen xitMDI LayoutMDI esize Resize ( Control) wDialog ( ) > 1 4 . 9 . , Form ti vate ( ) osed, Closing HChildActive , , . 5 . N E T . ) . 1, . : 2 (FormBorderStyle = FixedDialog); (MaximizeBox = False, MinimizedBox = False); 340 14. Windows , , Cancel, , (AcceptButton = __, Cancel Button = __1); DialogResult , . ShowModal, DialogResult, . 14.5. , . Cancel . . 14.6 , . (. 14.11) Dialog Exit Dialog , , , Cancel (. 14.12) . 1 4 . 1 1 . . 1 4 . 1 2 . , . Cancel, . * Project > Add Windows Form. , , < . 14.10. 341 1 4 . 1 0 . Text TextAli gn MiddleCenter Text Name btnOK DialogResult OK Text OK Name btnCancel DialogResult Cancel Text Cancel AcceptButton btnOK Cancel Button btnCancel FormBorderStyle FixedDialog MaximizeBox False MinimizeBox False StartPosition CenterParent textBoxl, . Text , Info. , : public class Form2 : Form { private Label private TextBox label 1: textBoxl: private Button btnOK: private Button btnCancel; private Container components = null; public string Info { get { return textBoxl.Text: } } } , , . 342 14. Windows (BorderStyle = FixedSingle). : private void menuIteml_Click( object sender, EventArgs e ) { Form2 f = new Form2(); // if ( f.ShowDialogO == DialogResult.OK ) // 1abel1.Text = f.Info; } private void menuItem2_Click( object sender, EventArgs e ) { CloseO; // } , , ShowDialog. Info, . , , set. Form . , , new . Show ShowDi al Form, . Hide. , Show Hide Visible . Hi de CI ose , , , CI ose . Application Application, System.Windows.Forms, , , . Application . 14.11. 1 4 . 1 1 . Application AddMessageFiIter, RemoveMessageFi Iter . , , 1 IMessageFiIter . [27]. 343 Application DoEvents Exit ExitThread Run CommonAppOataRegi stry CompanyName CurrentCulture CurrentInputlanguage ProductName ProductVersion StartupPath ApplicationExit Idle ThreadExit . , Appl i cati onExi t Appl i cati on ( , ), System.Reflection. Application, . 344 14. Windows , , Graphics, System.Drawing. . , Graphics PaintEventArgs, Paint, : private void Forml_Paint( object sender, PaintEventArgs e ) { Graphics g = e.Graphics; // } CreateGraphics, : Graphics g; g = this.CreateGraphicsO; - Image. : Bitmap bm = new Bitmap( "d:\\picture.bmp" ); Graphics g = Graphics.FromImage( bm ); Graphics , , . , , : Brush ; Font ; Col or . 14.6 , , . . 14.13. 1 4 . 6 . using System; using System.Drawing; using System.Windows.Forms; namespace WindowsApplicationl { public partial class Forml : Form { public Forml { InitializeComponent(); } private void Forml_Paint( object sender, PaintEventArgs e ) { using ( Graphics g = e.Graphics ) { // 1 345 using ( Pen pen = new Pen( Col or. Red ) ) 112 { g.DrawLine( pen. 0, 0, 200. 100 ); g.DrawEllipse( pen. new Rectangle(50, 50, 100, 150) ): } string s = "Sample Text"; Font font = new Font( "Arial", 18 ); SolidBrush brush = new SolidBrush( Col or. Black ); float x = 100.OF; float = 20.OF; g.DrawString( s, font, brush, x. ); font.DisposeO; brush.DisposeO: /7 3 //4 // 5 // . 1 4 . 1 3 . , Dispose. 1 # using : using ( ) , System. IDisposable, Dispose. , , , . Dispose , . using Dispose , . 1 2. 1 using # : . using 12. 346 14. Windows 3 4 , Di spose, 5 6. , , , -, , -, , -, . Windows- , : . Windows-: Microsoft, Apple, , , , : , , , , . . . . , , * . , , (, - ' ? ) , - . . Windows-. , . N E T : , , . , , < , ( , ] [17], [18], [20], [31]. : , i ! 1 5 # # Visual Studio: , X M L . # : A S P . N E T (- -) A D O . N E T ( ). , , + + , # , . X M L , , . , -, - # , - , , , , . . # : . , , . , , , , . (unsafe) . 348 15. # , C L R . unsafe, . unsafe , . , , , , . . , . , : public unsafe struct Node { public int Value; . public Node* Left; public Node* Right; } Node , Left Right. , : public struct Node { public int Value; public, unsafe Node* Left; public unsafe Node* Right; } unsafe : unsafe , , . , , ! /unsafe. Visu Studio (Project Properties Configuration Properties Build Allow Unsafe Cod* . : * ; , , . , . 6i , , , , * 349 : sbyte, byte, short, ushort, int, uint, long, uiong, char, float, double, decimal, bool void. , . voi d , , , (, ). voi d , , - , , . : int* // int Node* pNode; // Node void* // i n t * [ ] m; // int int** // int d; , : int* . , : // int . object, object . , . , . , , , . , , . . void. null. , : ( ). 350 15. # . , , 1 : 1. : : int = 5: int = &; // // , , , . : int* = ; , : i n t [ ] b = new i n t [ ] {10. 20. 30, 50}; // fixed ( int* t = b ) { . . . }; // fixed ( int* t = &b[0] ) { . . . } ; // f i xed . 2. : char* V = (char )0xl2F69E; 0xl2F69E , (char ) : char. , , . 3. : i nt* = nul 1 : 1 , (undefined behavior), . 351 4. : int* s = stackalloc int [ 1 0 ] ; stackalloc 10 int ( 10 ) s, . () # . , , ( . ). . . 15.1. 1 5 . 1 . , , -> [] & ++, -- +, - ==, ! = , < > , < = , > = , . stackalloc . , . , , , . , , : int = 5; int* = &; Console.WriteLineC * ): Console.WriteLineC ++() ) ; i n t [ ] b = new i n t [ ] {10, 20, 30, 50}; // // // // // , : 5 : 352 15. # // fixed ( int t = b ) // int* z = t; for (int i = 0; i < b.Length; ++i ) t[i] += 5; z += 5; ++z; // ( 5) // ( 5) // Console.WriteLineC &t[5] - t ); // fixed , , , , . , . : t z, . , . , . , . ( , , ) , . , $ , , . , . sizeof (), sizeof ( ). , , . - . , , , : short* ; ... ++; long* q; q++; // 2 // q 4 353 , . , 5. . . , ++ =10; , . , , , 10, , . : = 10; ++; ()++, , , . t: uint = 0xAB10234F; byte t = (byte)&x; for ( int i = 0; i < 4: ++i ) Console.Write("{0:X} ", t++ ) ; // : 4F 23 10 AB , t . 15.1 : 1 5 . 1 . using System; namespace ConsoleApplicationl { class A { public int value -. 20; struct { public int a; class Program { unsafe static void MainO { A n = new A O ; fixed ( int* pn = &n.value ) ++(pn); Console.WriteLineC "n = " + n.value ); b; pb = &b; // : 21 354 15. # 15.1 {) pb->a - 100; Console.WriteLineC b.a ); } // : 100 ' } } stackalloc : stackalloc [ ] . , System.StackOverflowException. , . int 0 4: int* = stackalloc int [5]; for int i = 0; i < 5: ++i ) { pC1]-1: Console.WriteC p[i] + " " ): } // : 0 1 2 3 4 15.2 , # . IntToString , . 1 5 . 2 . : using System: class Test { static string IntToString ( int value ) { int n - value >- 0 ? value : -value; unsafe { char* buffer = stackalloc char[16]; char* p = buffer + 16; do { --p = (charX n % 10 + '0' ); n /- 10; } while ( n !- 0 ); if ( value < 0 ) --p - '-': return new stringC p. 0, (int)C buffer + 16 - p ) ); } } static void MainO { Console.WriteLineC IntToStringC 12345 ) ); Console.WriteLineC IntToStringC -999 ) ); } } 355 : ; , ; . , , H T M L , . . N E T , System.Text.RegularExpressions. ' (), . : . , , . , , , , \d\d. . , IP-, , , . . .NET Perl 5. , . , , . , , , ! . 15.2 , . , . 15.3, , , , . , . 356 15. # 1 5 . 2 . , \ri . t cat, cut, clt, c{t . . [] , . c[aul]t cat, cut clt, c [ a - z ] t cat, cbt, cct, cdt,czt , , . c[aul]t cbt, c2t, cXt . ., c[a-zA-Z]t cnt, clt, c4t, c3t . . \w - , c\wt cat, cut, clt, ClOt . ., c{t, c;t . . \W - , , c\Wt c{t, c;t, t . ., cat, cut, clt, clOt . . \s , , (\t, \v), (\, \), ( \ f ) \s\w\w\w\s , \S , , \s\S\S\s , \d c\dt clt, c 2 t , c 9 t \D C\Dt clt, c2t c9t 1 5 . 3 . , , , , , , 357 . \Z , , \b , , ( , \w \W) , , , ~cat cat, , cat$ cat, ( ), $ , , . . , . , , \d\d\d\d\d \d{5}. 11111, 12345, 53332 . . . 15.4. 1 5 . 4 . cat ct, cat, caat, caaaaaaaaaaat . . ca+t cat, caat, caaaaaaaaaaat . . ca?t ct cat {} ca{3}t caaat, ( c a t ) { 2 } 1 catcat {".} { 3 , } t caaat, caaaat, caaaaaaaaaaaat . . {.} ca{2,4}t caat, caaat caaaat . 1 358 15. # . , , cat>, dog horse, cat\|dog\|horse , , . : (, ): t-+]?\d+ ( , ): C-+]?\d+\.?\d* (): [A-Z]\d{3}[A-Z]{2}\d\dRUS , - , . , \ . , \\. , cat.doc cat\.doc. , , ( !), 1 . . , , , ( ). , , , . : (?<_>_) , , . , , , nnn-rm-nn. : (?<num>\d\d\d-\d\d-\d\d) , , , .doc. 359 num . . , , , 1. , , . , (\w)M (wa//, mass, cooperate)1. , . : ()\s+()\s+(?<sum>\d+)\spy6\.\s+Hy . \1 , . . , , 5 . , 53459 . , IP-: ((\d{1.3}\.){3}\d{1.3}) , . . IP-: 212.46.197.69, 212.194.5.106, 209.122.173.160. , , . 1. , {3}. , , . , (?<s>\w)\k<s>, s , , \ . . , , . (?# ), , 2 # . .NET . N E T System.Text.RegularExpressions. 1 2 \w\w, - . RegexOptions. IgnorePatternWhitespace. 360 15. # Regex, . , . : Regex () ; Regex(String) ; RegexCString, RegexOptions) RegexOptions ( , ). , , , : Regex = new Regex( @"\b(?<word>\w+)\s+(\k<word>)\b". RegexOpti ons.IgnoreCase ); , , IsMatch, Match Matches. IsMatch true, , , , false . 15.3 . , , , . 1 5 . 3 . ( IsMatch) using System; usi ng System.Text.RegularExpressions; public class Test { public static void MainO { Regex r = new Regex( @"\b(?<word>\w+)[.,:;!? ]\s(\k<word>)\b", RegexOptions.IgnoreCase ) ; string t s t l = "Oh. oh! Give me more!"; if ( r.IsMatch( t s t l ) ) Console.WriteLine( " t s t l yes" ); else Console.WriteLine( " t s t l no" ); string tst2 = "Oh give me. give me more!"; if ( r.IsMatch( tst2 ) ) Console.WriteLineC " tst2 yes" ); else Console.WriteLine( " tst2 no" ); } } : t s t l yes tst2 no 361 tst2 , . , , - ( \ s ) ( . ) : Regex = new Regex( <a"\b(?<word>\w+)[..:;!? ].(\k<word>)\b". RegexOptions.IgnoreCase ) ; Match Regex, IsMatch, , , Match , . 15.4, . 1 5 . 4 . ( M a t c h ) using System; using System. Text.RegularExpressions; public class Test { public static void MainO { string text = " - $4, - $3, - $10."; string pattern = @"(\w+) - \ $ ( \ d + ) [ . . ] " ; Regex Match int total = new Regex( pattern ); = r.Match( text ); = 0; while ( m.Success ) { Console.WriteLine( m ); total += int.Parse( m.Groups[2].ToStringO ); m = m.NextMatchO; } Console.WriteLine( ": $" + total ); } } : - $4, - $3, - $10. : $17 Match , pattern. Match Groups, , . , , , , , . . , 362 15. # , , ; string pattern - @"(?'name'\w+) - \$(?'price'\d+).,]"; total += int.ParseC m.Groups["price"].ToString() ); NextMatch Match , . Matches Regex MatchCollection , . Split Regex. , , . 15.5 15.4 . 1 5 . 5 . ( Split) using System; usi ng System.Col 1ecti ons.Generi ; using System.Text.RegularExpressions; public class Test { public static void MainO { string text = " - $4, -$3. - $10."; string pattern = " [ - ..]+"; Regex r ' = new Regex( pattern ); List<string> words = new List<string>( r.SplitC text ) ); foreach ( string word in words ) Console.WriteLineC word ); ' ' ' i $4 $3 $10 Repl Regex . . , $ . .: string text = " - $4, -$3, - $10."; string textl = Regex.Repl( text, @"\$", "y.e." ); 363 MatchEvaluator, , . Regex Match System.Text.RegularExpressions , , Capture , ; CaptureCol 1 ecti on , ; Group Capture . . 8 -. , . : ppp-48.pool-113.spbnit.ru - - [31//2002:02:08:32 +0400] "GET / /1.1" .200 2434 "http://www.price.ru/bin/price/firmlnfo_f?fid=10922&where=01&base=2" "Mozilla/4.0 (compatible; MSIE 6.0; Windows NT 5.1)" 81.24.130.7 - - [31/May/2002:08:13:17 +0400] "GET /swf/menu.swf HTTP/1.~200 4682 "-" "Mozilla/4.0 (compatible; MSIE 5.01; Windows 98)" 81.24.130.7 - - [31/May/2002:08:13:17 +0400] "GET /swf/header.swf HTTP/1.1" 200 21244 "-" "Mozilla/4.0 (compatible; MSIE 5.01; Windows 98)" gate.solvo.ru - - [31/May/2002:10:43:03 +0400] "GET / HTTP/1.0" 200 2422 "http://www.price.ru/bin/price/firminfo_f?fid=10922&where0I&base=l" "Mozilla/4.0 (compatible; MSIE 5.0: Windows NT; DigExt)" , . , , 0 , , 5 (GET ), 8 (200 ) , , 9 . 15.6 H T M L , , , . 1 5 . 6 . - using System; using System.10; usi ng System.Col 1ecti ons.Generi ; using System.Text.RegularExpressions: public class Test { public static void MainO ^ & 364 15. # 15.6 () StreamReader f = new StreamReaderC "accessjog" ); StreamWriter w = new StreamWriterC "report.htm" ): Regex get Regex new RegexC "GET" ); new RegexC " " ); . 15.1. j?>block209-209-002. octetgroup.net j 162781 j J81.24.130.7 jgate. 174756 sohro. [212.113.108.164 ru " {113692} ~ ~ 12611991 . 1 5 . 1 . - a c c e s s j o g , , items. , , GET 2 0 0 5 8, 365 XML ( 9) - , , 0. H T M L - report.htm . , , Internet Explorer. , 1 .NET . XML X M L (extensible Markup Language) . , , , , : , , . . , . X M L : , . , , , : <summary> </summary> = , . X M L . X M L . , X M L , . .NET XML, System. Xml .NET , X M L . . . , . N E T XML-. , , / / / . , , ,- . . . 15.5. , Perl . 366 15. # 1 5 . 5 . <> <code> ( <1>) <example> <exception> <include file-'' path='nyTb[@name="Hfl"]" /> <Tist> <param> <paramref> <permission> <remarks> (, . .) <returns> <see cref=""> <seealso ^=""> <summary> (, . .) <value> : ///<summary> </summary> /7/<param name="i"> </> ///<seealso cref="System.Double"> double </seealso> ///<returns> </returns> ///<remarks> /// </remarks> public double Sin( double i ) { ... } 1 1 X M L /doc: _. xml. ^ Visual Studio (Project Properties) Build ( XML documentation file). . Visual Studio 2003 H T M L Tools Build Comment Web Pages. , , # : - -, , A D O . N E T . . , , . ADO.NET , . . N E T A D O . N E T (ActiveX D a t a Objects), . A D O . N E T : (content components) (managed- provider components). DataSet, . , DataTable, DataRow, DataColumn DataRelation. . DataSet DataTable DataRelation. DataSet , X M L , . ( ). Connection, Command DataReader. DataAdapter . , X M L - , , Excel. A D O . N E T : DataSet; - ; , DataSet, . A D O . N E T System.Data, System.Data.Common, System.Data.01 eDb, System.Data.SqlClient System.Data.SqlTypes. Visual Studio . N E T , . M S D E . Server Explorer (View Server Explorer) S Q L Server . , . ASP.NET A S P . N E T (Active Server Pages for . N E T ) - .NET, - -. , , 368 15. # . , , . : F T P - , - (Apache, IIS), , . - , W o r l d W i d e W e b . , . - H T T P / H T T P S . - : Apache ( - ; U n i x - ); IIS (Internet Information Services) Microsoft. IIS Microsoft , Win dows 2000 Professional . IIS Windows. aspnetregiis, . - , IIS , . - aspx. ASP.NET Web Application. , Location U R L - , . , W i n d o w s - , - Web Form Controls, H T M L , Windows Forms. - , W i n d o w s - : , . H T M L - aspx ( HTML ) # aspx.cs. , System.Web.UI.Page. (aspx-) . , A S P . N E T . Web Form Controls A S P . N E T , - . # , aspx-. , , . 369 ASP.NET , DataGrid DataList, , ( ADO DataSet). - . X M L , . - , H T T P S M T P . - . N E T .NET, IIS-. - , . N E T : , , , , . , . , , , - . , - ( , ) . - , XML-, H T T P . -, A S P . N E T , IIS-. - asmx. , aspx-, # asm.cs, -. , -, System.Web. Services.WebService. - ASP. NET Web Service. # , , .NET. , , [13], [20], [21], [23], [27], [ 3 1 ] - [ 3 3 ] . , , H T M L , X M L . ! , , . 1. : 1-3. . ( ). Math, # , . 64. , . , , . 1. zt = cos + sin + cos + sin ; z 2 = ^ - ^ s i n ^ - 8 j . 2. z, = cos + sin + cos + sin ; 2 = 2>/2 cos zx = sin 2 + sin5a - sin j 5. 6. 8. 2fll a\ -cos 4J 2 z2 = 4 cos cos^a cos 4a. - . a + ; 1 8 = 2 S I N A . z2 = cos a + cos a. Zj = cos a + cos 2a + cos 6a + cos 7a; 2(3 2 j - ._ ; cos a - cos 3a + cos 5a 2 , = l - - s i n 2 a + cos2a; 4 z<=cas,\-n z2=tg3a. zj= . z cosa + l - 2 s i n 2a sin2a + sin5a - sin3a 4. sm 4/ z, = cos x + sin + ^ sin 2x -1; = 2 sin. 2 z2 = sin(y + x) sin(y - 2. 371 2. : 4, . 1. , , . R . 372 374 376 2. , , , . . 3. 379 3. : 4, , . 1. , (. 1 2 ) , dx. , . . 2. , , 2 2. 3. , , # dx . . , . 380 381 4. ( lf 2 3 + 1 >-. 2 " 1 - 3 - 2 -1) 1 arcsinx = + > - - - = 2-4 - 2 n ( 2 n + 1) 21 19. - + 1-3- + 2-3 20. 1-3-5* + 2-4-5 1-3-5-7- + 2-4-6-7 . 2-4-6-8- <1. arccosx= 2 2-4--- 2 ( 2 + 1 ) 7 1- 1-3-5-* = \ + + + 2 \|^ 2-3 2-4-5 2-4-6-7 3 1-3-5-7-* + ... 2-4-6-8- <1- 4. : 4, , 5. , , : , , , . , , . . . 1 , , . . : , , . . , . 2 , , . . : , , 382 . . , . 3 , . , , . . . , . 4 , . . . , . 5 . , . , . , . ( ), , , . , . 7 , . , , , , , () . , . 8 . (, , ) 4. 383 . . , . , . 9 . (, , ) . . , . , . 10 2 + + . , : , -; , . 11 , . , , . . . , . 12 , . , , . . , . 13 , . . . , . 384 14 , . , , . , . 15 , , . . . . , . 16 , , . . . . , . 17 , - c o s ( x ) = 0. , (. for) . . , . 18 2 , + + = 0. , . . , . 19 , , , . . . . 5. 385 , . . . . , . 20 . . (, , ). . [0, 255]. . . , . 5. : 6, . 1 , , : ; , . 2 , , : ; , . 3 , , : 386 , . , , (, , ). 4 , , : ; , . , , . . 5 , , : ; , . , , [, ]. . 6 , , : ; , . , , , . 7 , , : , . , , , , . 1 5. 387 8 , , : ; , . , , , . 9 , , : ; , . , , , . 10 , , : ; , , . , , , , . 11 , , : ; , . , , [, ]. . 12 , , : ; , . 388 , , [, ], . 13 , , : , ; , . 14 , , : , ; , . 15 , , : , ; , . , , (, , ). 16 , , : ; , . 17 , , : ; , , . , , , . 6. 389 18 , , : , ; , . , , 20%, . 19 , , : ; , . 20 , , : , . , , , . 6. : 6, . 1 . : , ; , . 2 . , . . , . 390 3 . : , ; , . 4 . : , ; , . 5 . : , ; , . 6 . : , ; , ^ - j- . 7 8 x 8 , k- k- . , . 8 . , . , . 6. 391 9 A{j , i - l < k < i + l , j - 1 < l<j + 1, (k, I) , . 10 10. , . 10 , ( . 9). 10 10. , . 11 . . , . 12 , , . , . 13 ( ), . 14 N k : , , , ; . 392 15 . , . . , . 16 . , . 17 , , (2, 2), (3, 3) . ., . , . 18 . : , ; , . 19 . : , ; , . 20 . : , * ; 7. 393 , - t- ^'- . 7. : 6, . 1 , . 2 , , . 3 , , . 4 , , . 5 , , . 6 , , . 7 , , , . 8 , , , . 394 9 , , . 10 < , , . 11 , , . 12 , , , . 13 , , , . 14 , , 0 9 , , 15 , , , . 16 , , . 17 , , , . 8. 395 18 , . 19 , , , . 20 , , . 8. : 7. , , : , , ; , ; . , , . . . 1 (). : ; ; , . 2 . : 396 ; ; , . 3 , . : ; , -; , ; , . 4 , , , . , . 5 , . , , : , ; , . 6 . , - ( - 397 8. ', ), , , . , . 7 . , - (, , ), , . [ , . ( 8 ) . > , - (iep, , , ), , , . 1 , . 9 , ; . : , ( , ); (+=, -=); /; , . 10 , . : , ( , ); (=, /=); , . 398 11 , . : , (, , , , , , ); /; , . 12 , : , , . , . 13 . , , . , , . , , , . , . 14 . , , . , , . , . 15 , . . , , 6 . , . 16 , : 8. ( ); 399 . , . : ; , ; , ; , . . , . 17 , : , ; . , . : ; , ; , ; , ; , . , . 18 , : . , . : 400 , ; , ; , . . , . 19 , : , ; ( ). . , . : , ; , ; , ; , ; , . , . 20 - , . . . 9. : 8. (, ), . , 9. 401 . Equals, , . Mai , . . 1 Point (). ColoredPoint Line (). Line ColoredLine PolyLine ( ). : ; , ; 2 Vehicle ( ). Plane (), () Ship (). (, , . .) . , , . . 3 . : word . : Q ; , ; , ; ( ). _. , i. 402 , . - +, . - + , + , . , , . : 33il2 -700 +5i - 21 _ : 4 3. _. - +, . - + , + , . , , . . : , ; ( ); ( ). 5 3. _. ' 0' ' 1'. , , . . . 9. 403 _ : , ; ( ); 6 1. . : ( ); ; . : ; , 1 ; , . : ( ); ; . 2. , ( ), . . : ; , ; , ; , ( ). 3. , . , 404 (. ), . , . : , ( ); ; . : ( ); ; , ; , ; , ( ) ; , == . 4. , . : ; ; ; (), . : , ; , ; , . , pri vate. . , , , / . 5. , . . , : 405 10. 2- RS RST 10 12 RS 10 JK 11 10 10 JK 11 RS 12 12 RST 13 10 14 2- 10 15 10 16 JK 17 RS 10 18 19 JK 20 2- 10 10. : 9. 1 STUDENT, : ( ). , : , STUDENT ( ); , , , 4,0 ( , ). 406 2 STUDENT, : ( ). , : , STUDENT ( ); , 4 5 ( , ). 3 STUDENT, : ( ). , : , STUDENT ( ); , 2 ( , ). 4 AER0FL0T, : , : , AER0FL0T ( ); , , , ( , ). 5 AER0FL0T, : 10. 407 , : , AER0FL0T ( ); , , ( , ). WORKER, : , : , WORKER ( ); , , ( , ). 7 TRAIN, : , : , TRAIN ( ); , ( , ). 8 TRAIN, : 408 , : , TRAIN ( ); , , ( , ). 9 TRAIN, : , : , TRAIN ( ); , ( , ). 10 MARSH, : , : , MARSH ( ); , ( , ). 11 MARSH, : , : , MARSH ( ); , , ( , ). 1 10. 409 12 NOTE, : , ; ( ). , : , NOTE ( ); , ( , ). 13 NOTE, : , ; ( ). , : , NOTE ( ); , , ( , ). 14 NOTE, : , ; ( ). , : , NOTE ( ); , ( , ). 15 ZNAK, : , ; ( ). 410 , : , ZNAK ( ); , ( , ). 16 ZNAK, : , ; ( ). , : , ZNAK ( ); , , ( , ). 17 ZNAK, : , ; ( ). , : , ZNAK ( ); , , ( , ). 18 PRICE, : , ; , : , PRICE ( ); 1 1 . 411 , ( , ). 19 PRICE, : , ; , : , PRICE ( ); , , ( , ). 20 ORDER, : , : , ORDER ( ); , , ( , ). 1 1 . : 9, 13. 9, . IComparable - . 412 12. Windows- : 14. 1. : Windows-, . . ., . , - 1 Input, Calc Exit. Input , : TextBox ; ( ) CheckBox; Button. : / . Calc . Exit . 2 Size, Color, Paint, Quit. Paint . Quit . Size , : TextBox ; (Red, Green, Blue) CheckBox; Button. Paint . Paint , . 12. Windows- 413 3 Input, Work, Exit. Exit . Input , : TextBox Radius, Height, Density; (Volume, Mass) CheckBox; Button. , ; : / . Work . 4 Input, Calc, Draw, Exit. Exit . Input , : TextBox Radius; (Square, Length) CheckBox; Button. ; : (Square) / (Length). Calc . Draw , ( ). 5 input, Calc, About. About . Input , : TextBox Number 1, Number 2, Number 3; (Summ, Least multiple) CheckBox; Button. : (Summ) / (Least multiple). Calc . 414 6 Input, Calc, Quit. Calc . Quit . Input , : TextBox Number 1, Number 2; (Summa, Max divisor, Multiply) CheckBox; Button. ; ( , , ). Calc . 7 Begin, Help, About. About . Begin , : TextBox input; Label ; (2, 8, 16) RadioButton; Button Do . input; : , . Do . 8 Input color, Change, Exit, Help. Exit . Input color , : TextBox Red, Green, Blue; (Left, Right) CheckBox; Button. RGB- . Change (, ). 12. Windows- 415 9 Input size, Choose, Change, Exit. Exit . Change . Input size , : TextBox Size , Size ; Button. Choose , : (Increase, Decrease) RadloButton; Button. Size Size . , ( ). Change . . 10 Begin, Work, About. About . Begin , : TextBox Input word; (Upper case, Lower case) RadioButton; Button. . Work . 11 Input color, Change, Clear. Input color , : (Up, Down) CheckBox; (Red, Green, Blue) RadioButton; Button. ; , , : , . Change ( , ). Clear . 416 12 Translate, Help, About, Exit. Exit . Translate , : TextBox Binary number; TextBox (read-only); (8, 10, 16) RadioButton; Do Button. Binary number; : , . Do . 13 Reverse, About, Exit. About . Reverse , : TextBox Input; (Upper case, Reverse) CheckBox; Button. : / . . 14 Input, Show Exit. Exit . Input : 12. Windows- 417 length koef: , , / . Show . 15 Input, About Exit. Exit . About . Input : . Euro $ . 16 Begin, Work, About. About . Begin , : TextBox; (First letter, All letters) RadioButton; Button. : (First letter), (All letters). Work . 17 , , . 418 OpenFileDialog. . , . 18 H T M L . : HTML OpenFi leDialog. SaveFileDlalog. Memo. H T M L :  ; <BR>; < <; > &gt:; & &;; " ( ) Squot;. 19 Input, Draw, Clear. Input , : (Red, Green, Blue) RadioButton; Button. 12. Windows- 419 D r a w , . Clear . 20 Input, Change, Exit. Exit . C h a n g e . 100 100 . Input , : TextBox Size , Size ; (Increase, Decrease) RadioButton; Button. Size Size . , , ( ). C h a n g e . . , . 21 Windows-, . Choose, S h o w , Quit. S h o w . Quit . . . , (price) (hard drive). C h o o s e , : TextBox ; TextBox ; (Hard drive, Price) RadioButton; O K , Cancel Button. S h o w . S h o w , , . 422 ( ). Animate. Animate . . . . . 5 Windows- = - + - + -log2 . Open, Coefficients, Show, About, Exit. Exit . About . ( Open) OpenFileDialog. () . Coefficients ,. Show . . . # . (currency) . ( $ ) . , NumberFormatlnfo. , D d . , . , , 12 D3 012 , d.ddd..E+ddd d.ddd...e+ddd. , . 3 F f . , , , 0,12, F3, 0,120 G g . , . , ; single 7 , byte sbyte 3, decimal 29 & 424 . # [) d,ddd,ddd.ddd..., , . , (, 100, %) . , . X, using System; namespace ConsoleApplicationl { class Classl { static void MainO { int i = 1234; Console.WriteLineC i.ToStringC "C" ) ); Console.WriteLineC i.ToStringC "D5" ) ); Console.WriteLineC i.ToStringC "E" ). ); Console.WriteLineC i.ToStringC "G" ) ); Console.WriteLineC "{0,9:n2}", i ) ; Console.WriteLineC "{0,l:p3}", Console.WriteLineC "{0.1:x}", i i ); ); } } } : 1 234.00. 01234 1.234000+003 1234 1 234,00 123 400,000% 4d2 . 146. 1. . . # . - .: - - - . , 2006. - 488 . 2. . -, . - . : -, 2000. - 304 . 3. . #. - . : , 2004. - 880 . 4. . . - : , 2001. 352 . 5. . .NET Java-. - .: , 2003. 336 . 6. . . ++. . - .: , 1996. - 272 . 7. . # . . - .: , 2001. 304 . 8. . Visual + + .NET. - .: ; : B H V , 2004. - 528 . 9. . # . - .: -, 2003. - 688 . 10. . # . : . : , 2002. - 656 . 11. . # Builder. . .: , 2005. - 384 . 12. . # . : . : , 2002. - 656 . 13. . A S P . N E T # . - . : , 2003. - 304 . 14. . . C / C + + . : . - .: , 2001. - 464 . 15. . . . : . - .: , 2003. - 393 . 426 16. ., . : Visual + + .NET. .: , 2002. - 816 . 17. . MS Windows # . . 1. . : - , 2002. 576 . 18. . MS Windows # . . 2. . : - , 2002. 624 . 19. . . + + / # Visual Studio . N E T 2003. . .: -, 2004. 352 . 20. ., . Visual C # . N E T . . : , 2004. - 960 . 21. , ., . . # . 2 . .: , 2003. - 512 . 22. . # . .: , 2002. 352 . 23. . + + # . . .: , 2003. - 608 . 24. . # . "". .: -, 2001. - 576 . 25. . # . : , 2003. - 528 . 26. ., X. . .NET. . .: -, 2003. 336 . 27. . # .NET. . .: , 2002. - 796 . 28. . ., . . # : . .: - , 2003. - 560 . 29. . # : . - .: , 2002. - 512 : . 30. . # . . : , 2004. - 752 . 31. Microsoft Corporation. Windows- Microsoft Visual Basic . N E T Microsoft Visual C# .NET. . 70-306, 70-316. . : - , 2003. - 512 . 32. Microsoft Corporation. Web- Microsoft Visual Basic .NET Microsoft Visual C # .NET. M C A D / M C S D . - M.: - , 2003. 704 . 33. Microsoft Corporation. Web- X M L Microsoft Visual Basic . N E T Microsoft Visual C# .NET. M C A D / M C S D . M.: - , 2004. - 576 . !, , 43, 49 I-, , 43, 54 #define, , 288 #elif, , 288 #else, , 288 #endif, , 288 #endregion, , 288 #, , 288 #if, , 288 #line, , 288 #pragma, , 288 #region, , 288 #warning, , 288 %, , 43, 51 % = , , 43 &&, , 43, 56 &, , 43, 55 &=, , 44 ( ) , , 42 , ,. 43, 50 * = , , 43, 57 , , 42 -, , 43 --, , 42, 43, 47 -, , 48, 53 ., , 42 /, , 43, 50 / = , , 43 ?, , 43, 56 ??, , 310 @, , 30 , , 43, 55 = , , 44 \|, , 43, 55 \|, , 43, 56 \| = , , 44 -, , 43, 49 +, , 43, 52 + + , , 42, 43, 47 + = , , 43, 57 <, , 43, 54 , , 43, 5 4 = , , 4 4 < = , , 43, 54 >, , 43, 54 , , 43, 5 4 = , , 4 4 > = , , 43, 54 =, , 43, 57 - = , , 44, 57 , , 43, 54 Abort, , 240 Abs, , 65 abstract, , 101,181 Component, , 337 Concat, , 144 Console, , 19, 59, 262 const, , 41 ContainerControl, , 338 ContextMenu, , 331 Acos, , 65 ADO.NET, 367 ANSI, 22 API, 312 Append, , 147 AppendFormat, , 147 Application, , 342 Array, , 133 ArrayList, , 296 as, , 43, 194 Asin, , 65 ASP.NET, 367 AsyncCallback, , 242, 255 Atan, , 65 Atan2, , 65 base, , 117, 175 Beginlnvoke, , 242 BeginRead, , 250, 253 BeginWrite, , 250 BigMul, , 65 BinaryFormatter, , 268 Binary Reader, , 261 Binary Search, , 134 BinaryWriter, , 260 break, , 84 Brush, , 344 Button, , 326 Capture, , 363 case, , 74 catch, , 90 Ceiling, , 65 char, , 139 CheckBox; , 331 checked, , 42, 46 class, , 100, 305 Close, , 250, 255 CLR, 9 CLS, 14 Exception, , 89, 94 , , 65 explicit, , 168 Color, , 344 Combine, , 222 Compare, , 143 CompareOrdinal, , 144 CompareTo, , 144 File, , 263 FileAccess, , 249 FileAttributes, , 265 Filelnfo, , 263 continue, , 86 Control, , 323, 338 Convert, , 61 , , 134, 144 Cos, , 65 Cosh, , 65 D DataSet, , 367 decimal, , 34 default, , 74, 306 delegate, , 220 Deserialize, , 268 DialogResult, , 327 Dictionary<T,K>, , 302 Directory, , 263 Directorylnfo, , 263 Dispose, , 345 DivRem, , 65 DLL, 312 do while, , 79 , , 65 Endlnvoke, , 242 EndRead, , 250, 253 EndWrite, , 250 Enum, , 218 enum, , 215 Epsilon, , 73 Equals, , 184 8-, 24,29 event, , 233 EventArgs, , 235 EventHandler, , 235 428 FileMode, , 249 FileShare, , 249 FileStream, , 250 FileSystemlnfo, , 263 finally, , 90 FindMembers, , 280 fixed, , 352 Floor, , 65 Flush, , 250, 255 Font, , 344 for, , 81 foreach, , 136 Form, , 337, 338 Format, , 144 G GAC, 275 get, , 121 GetConstructors, , 280 GetEnumerator, , 207 GetEvents, , 280 GetFields, , 280 GetHashCode, , 184 Getlnterfaces, , 280 GetMembers, , 280 GetMethods, , 280 GetName, , 218 GetNestedTypes, , 280 GetNumericValue, , 140 GetProperties, , 280 GetType, , 184,280 global, , 287 goto, , 83 Graphics, , 344 Group, , 363 GroupBox, , 332 IAsyncResult, , 255 ICloneable, , 205 IComparable, , 199 IComparer, , 200 IDisposable, , 345 IEEERemainder, , 65 IEnumerable, , 198, 207, 219 IEnumerator, , 198, 207, 219 if, , 70 IIS, 368 ILDasm, , 273 implicit, , 168 IndexOf, , 134 Insert, , 144 internal, , 101 is, , 43, 194 IsControl, , 140 IsDigit, , 140 IsLetter, , 140 IsLetterOrDigit, , 140 IsLower, , 140 IsMatch, , 360 IsNumber, , 140 IsPunctuation, , 140 IsSeparator, , 140 IsUpper, , 140 IsWhiteSpace, , 140 JIT-, 9 Join, , 144 L N Label, , 326 LastlndexOf, , 134 Length, , 144 List<T>, , 300 ListBox, , 334 lock, , 242 Log, , 65 Log 10, , 66 Main, , 156 MainMenu, , 330 MarshalByRefObject, , 337 Match, , 361 Match, , 361 MatchCollection, , 362 Matches, , 362 MatchEvaluator, , 363 , , 66 MaxValue, , 34, 140 MDI, 337 MemberwiseClone, , 205 Menultem, , 330 MessageBox, , 322 Min, , 66 MinValue, , 34, 140 Monster, , 119 MSIL, 9 NaN, , 34 new , 175, 305 , 42, 48, 102 , 101 Next, , 149 NextBytes, , 149 NextDouble, , 149 NextMatch, , 362 null, , 30 nullable, , 36,309 O object, , 13, 183 OLE, 312 operator, , 161 out, , 113 override, , 179 params, , 154 Parse, , 61, 140 partial, , 308 Peek, , 256 , , 344 PI, 66 Pow, , 66 private, , 101 protected, , 101 public, , 101 RadioButton, , 331 Random, , 148 Read, , 62, 250 ReadBlock, , 256 ReadByte, , 250 ReadLine, , 62 readonly, , 105 ReadToEnd, , 256 ref, , 112 ReferenceEquals, , 184 Regex, , 360 RegexOptions, , 360 Remove, , 144, 225 Replace, , 144, 362 return, , 87 Reverse, , 134 Round, , 66 Run, , 316 S sealed, , 101,182 Seek, , 250 Serialize, , 268 set, , 121 Sign, , 66 Sin, , 66 Sinh, , 66 sizeof, , 352 Sleep, , 240 Sort, , 134 Split, , 144,303,362 Sqrt, , 66 stackalloc, , 354 Start, , 241 static , 118 , 101 Stream, , 250 StreamReader, , 255 Stream Writer, , 255 string, , 143 StringBuilder, , 147, 151 strong name, 275 struct, , 212, 305 429 Substring, , 144 switch, , 73 System. Collections, , 295 System.Collections.Generic, , 295, 300 System.Collections.Specialized, , 295 System.Drawing, , 344 System.Reflection, , 279 System.Text. RegularExpression, , 355 System.Windows.Forms, , 315 Tan, , 66 Tanh, , 66 TextBox, , 327 TextReader, , 255 TextWriter, , 255 this, , 114 Thread, , 239 ThreadStart, , 239 throw, , 93 ToCharArray, , 144 ToLower, , 140 ToString, , 60, 184 ToUpper, , 140 try, , 90 , , 280 typeof, , 42 unchecked, , 42, 46 Unicode, , 22, 24 unsafe , 348 , 348 using , 286 , 345 value, , 122 virtual, , 178 178 void, , 109 volatile, , 105 where, , 305 while, , 76 Write, , 60, 250 WriteByte, , 250 WriteLine, , 19, 60 XML, , 11 , 294 , 181 , 22 , 22 , 24 , 24 , 228 , 32 , 48 -, 253 , 242 , 293 , 283 , 87 , 10 , 215 , 182 , 312 , 10 , 164 , 292 , 40,70 , 62,246 , 246 -, 368 -, 368 -, 369 , 274 , 274 , 87 , 33 DOS-, 312 , 368 , 178 , 317 , 182 V-Z VMT, 365 yield, , 208 , 101 , 169 , 32 , 239 , 246 , 23, 42, 70, 355 , , 53 - , 15, 330 , 337 , 275 , 294 , 102 , 102 , 248, 260, 271 , 291 , 291 , 47 , 220 , 242 , 221 , 220 , 226 , 225 , 182 , 50 , 292 , 293 , 267 , 169 , 326, 337 , 312 , 23, 287 XML, 365 , 237 , 32 , 274 , 30 , 248, 271 , 248 , 248 , 337 , 106 , 314 , 117 , 246 , 274 , 25, 42 , 48 , 35 , 274 , 24 , 24 , 12 , 284 , 24 , 100 , 19 , 275 430 , 126 , 157, 161 , 116 , 39 , 12 , 47 , 305 , 12, 124, 188 , 337 , 307 , 274 , 274 , 46, 89 , 89 , 223 , 207 , 75 , 11 , 368 , 368 , 13, 100 , 181 , 182 , 101, 169 , 172 , 172 , 299 ' , 118 , 205 , 292 , 25 , 326 , 32 , 121, 158 , 347 , 22 , 295 , 300 , 294 , 292 , 15 , 23, 359 , 30 , 30 , 8 , 314, 325 , 367 , 367 , 59 , 15 , 15 , 26, 42 , 114 , 117 , 117 , 174 , 48,115 , 117 , 117 , 295 , 331 , 292 , 9 - , 23 , 24 , 59 , 293 , 26 , 30 , 29 , 49 , 39 , 273 , 28 , 126, 291 , 293 , 129 , 156 , 128 , 130 , 127 , 126 , 132 , 330 , 331 , 9, 273 , 355 , 84 , 18, 106 , 228 , 178 , 102 , 306 , 152 , 182 , 153 , 102 , 226 , 102 , 337 , 311 , 161 , 30 , 294 , 337 , 326 , 182 , 223 , 12, 172 , 347 , 348 , 143 , 337 , 45 , 274 , 274 , 274 , 25 , 40 , 111 , 309 , 306 , 8 , 233 , 91 , 313 , 359 , 226 , 14 , 9 , 11 , 101 , 211 - , 69 , 161 , 305 , 128 , 291 , 291 , 30 , 311 , 337 , 326, 337 , 337 , 337 , 337 , 17 , 337 , 16 , 322 , 16 431 , 42 , 24 break, 84 continue, 86 lock, 242 , 83 , 70 , 87 , 73 , 136 , 83 , 70 , 70 , 75 , 42 , 164 , 354 , 53 , 47 , 50 , 105, 353 , 47 , 161 , 56 , 49 , 310 , 51 , 54 , 161 , 55 , 49 , 49, 168 , 57 , 351 , 351 , 54 , 52 , 48 , 50 , 162 , 48 , 56 , 18, 100 , 274 , 51 , 273 , 232 , 48 , 49 , 49 , 292 , 15 , 113 , 220 , 111 , 284 , 156 , 107 , 284 , 112 , 76 , 300 , 307 , 223 , 274 , 238 , 60 , 152 , 161 , , 110 , 110 , 38 , 39 , 215 % , 211 , 215 , 8 , 205 , 357 , 86 , 178 , 284 , 39, 104 , 13,153,180,188 , 182 , 232 , 55 , 49 , 28 , 248, 271 , 311 -, 246 , 239 , 238 , 238 , 172 , 24 , 172 , 45, 49 , 11 , 44 , 57 , 57 , 214 , 23 , 47 , 59 , 69 , 69 , 312 , 14 , 248 , 273 , 57 , 13, 285 , 299 , 237 , 130 , 287 , 286 , 239 , 70 , 312 , 351 , 9 , 26 , 127 , 152 , 351 , 177 , 36, 214 , 93 , 225 , 355 , 8 , 249 , 249 , 249 , 153 , 153 , 274, 345 , 279 , 15 , 337 , 9, 272 , 273 , 273 432 , 169 , 120 , 178 , 177 , 54 , 283 , 367 , 267 , 108 , 275 , 23 , 23 , 17 , 21 , 24 , 304 , 87 , 293 , 25 , 52 , 232,312 , 233 , 233 , 312 , 45, 350 , 313 , 306 , 70 , 220 , 101 , 157 , 101 , 215 , 233 , 212 , 146 , 291 , 291 , 292 , 291 , 291 , 9 , 8 , 35 , 118 , 117 , 102 , 102 , 31, 292 , 15 , 143 , 143 , 29 , 212 , 294 , 69 , 132 , 76 , 178 , 365 , 169 , 189 , 100 , 19,106 , 161 , 215 , 212 , 75, 137 , 32 , 33 , 169 , 32 , 31 , 220 , 35 , 19 , 143 , 309 , 215 , 304 , 233 , 35 , 32 , 308, 310 , 284 , 348 , 50 , 162 , 48 , 226 , 36, 214 , 312 , 29 , 56 , 248,260,271 , 368 , 226 , 19 , 161 , 294 , 19 -, 294 -, 293 -, 294 , 31 , 32 , 87 , 313 , 82 , 81 , 79 , 76 , 308, 310 , 273 , 246 , 224 , 146 , 13, 101 , 102 , 325 , 22 , 273 \| - , (60), . . # 2.0 ( 2 0 0 5 ) , , , . # , BASIC + + , . 20 . - ^ & ^ h t t p : / / i p s . i f m o . r u . , : .NET; #\|\| - ; .NET; , , , , , , ; - , ), , " : 197198, -, / 619 .: (812) 703-73-74, postbook@piter.com 61093, -93, / 9130 .: (057) 712-27-05, piter@kharkov.piter.co w w w . p i t e r . c o m Вам также может понравиться
Benefits of walking in the morning Walking benefits Why we should walk in the morning 30 minutes of walking can change your life, walking in the morning is beneficial for your health and reduce the diseases chance like heart problem, blood pressure, diabetes, and reduce obesity, walking in the morning offers you mind and body added benefits, there is the reason behind you should walk at the morning. 1. In the morning air is less polluted and we get fresh air which goes to our mind and the body makes us feel happy and energetic. 2. In the morning temperature is cool which is good for our body and our body gets a pleasant environment. 3. There is less traffic chock in the morning. Let’s see what the science says about the morning walk. Increase your metabolism running Researcher shows that walking at any time of the day is beneficial for health. you might be feeling lazy for a morning walk, you just think walking is just walking but in reality that seems different, walking at different times has different benefits. Therefore, working cardio with a walk would enhance your metabolism. The term metabolism is commonly used to refer specifically to the breakdown of food and its transformation into energy for the entire day and keep you more energetic and active to do any work Prevent daily life disease walking benefits According to the researcher, morning walks are extremely beneficial in preventing or reducing the trades of daily life diseases like stress, anxiety, diabetes, thyroid, hypertension, and health-related problems. Keep sugar levels under control morning walk control sugar According to the data about 422 million people worldwide have diabetes. Diabetes is a long-standing, anabolism disease distinguished by raised levels of blood glucose, which show over time to serious injury to the heart, blood vessels, and nerves. Type 2 diabetes, is usually common in adults and occurs when your body doesn’t make enough insulin. In the past decades, the generality of type 2 diabetes has risen across countries. Type 1 diabetes, once known as insulin-dependent diabetes, is a long-standing condition in which the pancreas produces little or no insulin by itself. There is a globally agreed target to stop the rise in obesity and diabetes by 2025 We Could control sugar level by walking for 30 minutes every day, when we walk our blood start circulating fast and cell start utilizing glucose in the bloodstream more efficiently and start reducing fat from your body which is really helpful to reduce blood sugar. Read more that food help in diabetes. Melt body fats burn belly fat However, we do heavy workouts at the gym but the next morning we feel lazy and tired to go to the gym the morning walk seems easy than a heavy gym workout. Although when we walk outside we see nature and get the fresh air, our mind feels happy but in the gym, we don’t get these things. Walking is very helpful in burning the body fats and it shows effective results when we walk at least 30 minutes daily. It’s very helpful to burn the belly fats when we walk daily fats start converted into energy and it helps you to get back in your shape. Improve mental health When we do a morning walk daily we feel happy and keep ourselves positive all day, endorphins hormone is released when we walk which is made by the pituitary gland and central nervous system this hormone makes us happy and stress-free. • Morning walks release stress, depression, and anxiety, Most youth and adults are surviving with depression, and morning walk cures these things definitely. • Morning walk improves the memory and helps to improve cognitive skills, morning walk is very good for old age people because when you start getting old your memory starts losing, so it is very helpful for those people who are more than 60 years old Control cholesterol level We know very well we need a certain amount of cholesterol in our body to maintain optimum health and build cell membranes. However, when the cholesterol increases there are more chances you may experience heart problems when there is an excessive amount of blood lipids, especially when they are in the form of LDL cholesterol. Improve lung capacity When you walk you need extra oxygen to maintain an oxidation reaction quotient in the cells of your body, therefore these reactions cause a high demand for oxygen supply in your body that makes the lungs pump extra oxygen. Hence this, in turn, helps the lungs improve their capacity. Boost your immune system When you do walk blood starts circulating fast in your body to provide the proper energy, walking improves the blood circulation system in your body. This has wonderful effects on the immune system. Moreover, it also increases the supply of oxygen throughout the body. walking for just 30 minutes a day to improve your immune system and protects you from various diseases and illnesses. Make your skin glow According to the researcher that any exercise which increases blood circulation gives you a healthy skin glow. There is no better exercise than walking. walking improves blood circulation due to this it prevents pimples, acne, wrinkles, and other skin problems. Only 30 minutes of walking daily give you an extra glow on your face.
1. Laura Nenzi 2. jSSTL Overview HTTPS SSH jSSTL : java Signal Spatio Temporal Logic jSSTL is a Java library for the specification and verification of spatio-temporal properties of dynamical systems described as spatio-temporal traces over a weighted graph. Given a trace x(t,l), a weighted graph and a formula Phi, it returns the boolean and quantitative spatio-temporal signals of the satisfaction at each time and in each location. It consists of three main packages: core, util and io. • core.formula contains the classes used to specify an SSTL formulas. They mimic the abstract syntax tree of formulas. core.monitor provides the implementation of the monitoring algorithms to verify an SSTL formulas. The monitoring is performed via a visit of a formula that implements a bottom-up evaluation. core.space provides the classes for the representation of the graph. It relies on JGraphT, a free Java graph library that provides mathematical graph-theory objects and algorithms. The methods booleanCheck(graphModel, inputSignal) and quantitativeCheck(graphModel, inputSignal) of a formula produce the boolean and quantitative signals for the trace inputSignal with a topology of the considered locations as a graphModel. • util contains the classes used to represent and manage the signals. • io provides a set of classes that can be used to read graph models and input signals from an input stream and to write monitoring results to an output stream. Currently, CSV and tabular based ascii files are supported for both input and output of signals. An Eclipse plug-in providing front-end for SSTL monitoring is currently under development. How to run A running example is provided within the library. Who do I talk to? • laura.nenzi-at-imtlucca.it • michele.loreti-at-unifi.it
ShellShocker is now owned by minttm.com Shellshocker.net is now taken over by MintTM(https://www.minttm.com) a leading website development company empowering startups to build the foundation of their dream ideas. What is #shellshock? Shellshock (CVE-2014-6271, CVE-2014-6277, CVE-2014-6278, CVE-2014-7169, CVE-2014-7186, CVE-2014-7187) is a vulnerability in GNU's bash shell that gives attackers access to run remote commands on a vulnerable system. If your system has not updated bash in since Tue Sep 30 2014: 1:32PM EST (See patch history), you're most definitely vulnerable and have been since first boot. This security vulnerability affects versions 1.14 (released in 1994) to the most recent version 4.3 according to NVD. You can use this website to get information on how to identify the vulnerability and the material to resolve it. This project can be found at Github. Check it out here. Stay safe! Testing Your System To test your system, you can run shellshock_test.sh. You can view the source and download it here - shellshock_test.sh on GitHub. If you want to test each exploit individually without running the script above, feel free! They are listed below. Exploit 1 (CVE-2014-6271) There are a few different ways to test if your system is vulnerable to shellshock. Try running the following command in a shell. env x='() { :;}; echo vulnerable' bash -c "echo this is a test" If you see "vulnerable" you need to update bash. Otherwise, you should be good to go. Exploit 2 (CVE-2014-7169) Even after upgrading bash you may still be vulnerable to this exploit. Try running the following code. env X='() { (shellshocker.net)=>\' bash -c "echo date"; cat echo; rm ./echo If the above command outputs the current date (it may also show errors), you are still vulnerable. Exploit 3 (???) Here is another variation of the exploit. Please leave a comment below if you know the CVE of this exploit. env X=' () { }; echo hello' bash -c 'date' If the above command outputs "hello", you are vulnerable. Exploit 4 (CVE-2014-7186) bash -c 'true <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF <<EOF' \|\| echo "CVE-2014-7186 vulnerable, redir_stack" A vulnerable system will echo the text "CVE-2014-7186 vulnerable, redir_stack". Exploit 5 (CVE-2014-7187) (for x in {1..200} ; do echo "for x$x in ; do :"; done; for x in {1..200} ; do echo done ; done) \| bash \|\| echo "CVE-2014-7187 vulnerable, word_lineno" A vulnerable system will echo the text "CVE-2014-7187 vulnerable, word_lineno". Exploit 6 (CVE-2014-6278) shellshocker='() { echo You are vulnerable; }' bash -c shellshocker You shouldn't see "You are vulnerable", if you're patched you will see "bash: shellshocker: command not found" Exploit 7 (CVE-2014-6277) bash -c "f() { x() { _;}; x() { _;} <<a; }" 2>/dev/null \|\| echo vulnerable If the command outputs "vulnerable", you are vulnerable. If you've tested your system, please leave a comment below. Don't forget to include your bash version and what OS you're running. Type bash --version for bash, and cat /etc/release for your OS. How to fix ShellShock CentOS, Ubuntu, Linux systems Shellshock is a vulnerability in bash. In order to patch your vulnerable system, you will need to get the most up to date version of bash available from GNU.org. Depending on your package manager (yum, apt-get, etc) you may be able to just run a yum update and you'll be good to go. Here's how that's done: yum update bash -y For Ubuntu Systems: apt-get update; apt-get install --only-upgrade bash For Arch Linux: pacman -Syu If your package manager doesn't find an update, you will need to build bash from src. Building From Source You can patch bash with one command using our bash patcher, just run the following command and you should be good to go! Make sure you have patch installed before you run this command. sudo apt-get install patch (yum install patch) etc... curl https://shellshocker.net/fixbash \| sh If you want to do it yourself, feel free. Here are all the commands you'll need. cd ~/ mkdir bash cd bash wget https://ftp.gnu.org/gnu/bash/bash-4.3.tar.gz #download all patches while [ true ]; do i=`expr $i + 1`; wget -N https://ftp.gnu.org/gnu/bash/bash-4.3-patches/bash43-$(printf '%03g' $i); if [ $? -ne 0 ]; then break; fi; done tar zxvf bash-4.3.tar.gz cd bash-4.3 for p in `ls ../bash43-[0-9][0-9][0-9]`; do patch -p0 < $p; done ./configure && make && make install OS X If you're running OS X, Apple has released official patches for Mavericks, Mountain Lion and Lion. You can also download and compile bash yourself using brew or MacPorts. We recommend using brew - Go to http://brew.sh/ and install brew on your system. Once you have brew installed, run the following commands to update your system brew update brew install bash sudo sh -c 'echo "/usr/local/bin/bash" >> /etc/shells' chsh -s /usr/local/bin/bash sudo mv /bin/bash /bin/bash-backup sudo ln -s /usr/local/bin/bash /bin/bash If you're using MacPorts, run the following: sudo port selfupdate sudo port upgrade bash Once you've updated, try the exploit again and report back your findings.
Merge branch 'upstream_libresoc' into libresoc_master [c4m-jtag.git] / setup.py 1 from setuptools import setup, find_packages 2 3 4 def scm_version(): 5 def local_scheme(version): 6 if version.tag and not version.distance: 7 return version.format_with("") 8 else: 9 return version.format_choice("+{node}", "+{node}.dirty") 10 return { 11 "relative_to": __file__, 12 "version_scheme": "guess-next-dev", 13 "local_scheme": local_scheme 14 } 15 16 17 setup( 18 name="c4m_jtag", 19 use_scm_version=scm_version(), 20 author="Staf Verhaegen", 21 author_email="staf@fibraservi.eu", 22 description="", 23 license="multi", 24 python_requires="~=3.6", 25 setup_requires=["setuptools_scm"], 26 27 # removing cocotb, causing unnecessary dependency and install problems 28 install_requires=["setuptools", "nmigen", "nmigen-soc", "modgrammar"], 29 30 # unit tests require cocotb: main operation does not 31 tests_require=['cocotb'], 32 33 include_package_data=True, 34 packages=find_packages(), 35 project_urls={ 36 #"Documentation": "???", 37 "Source Code": "https://gitlab.com/Chips4Makers/c4m-jtag", 38 "Bug Tracker": "https://gitlab.com/Chips4Makers/c4m-jtag/issues", 39 }, 40 )
Probabilistic Prefetching Scheme for P2P VoD Applications With Frequent Seeks Source: Ryefone Inc. Favorite Free registration required In Peer-to-Peer Video-on-Demand (P2P VoD) applications, users tend to seek to the positions that they are interested in. The frequent seeks raise a great challenge to the design of the prefetching scheme. In this paper, the authors propose a probabilistic prefetching framework to reduce the seeking distance. Each peer performs prefetching based on the segment access probability, which is estimated from the seeking statistics in the previous sessions. It is a challenging task to collect the seeking statistics in a distributed P2P network. In the proposed framework, they employ FM sketches to represent the seeking statistics, thus greatly reducing the space and time complexity. Format:PDF Size:235.64 Date:Jan 2008
Combining Inferences from Imputed Data Sets With imputations, different sets of the point and variance estimates for a parameter can be computed. Suppose that and are the point and variance estimates, respectively, from the th imputed data set, = 1, 2, ..., . Then the combined point estimate for from multiple imputation is the average of the complete-data estimates: Suppose that is the within-imputation variance, which is the average of the complete-data estimates: And suppose that is the between-imputation variance: Then the variance estimate associated with is the total variance (Rubin 1987) The statistic is approximately distributed as with degrees of freedom (Rubin 1987), where The degrees of freedom depend on and the ratio The ratio is called the relative increase in variance due to nonresponse (Rubin 1987). When there is no missing information about , the values of and are both zero. With a large value of or a small value of , the degrees of freedom will be large and the distribution of will be approximately normal. Another useful statistic is the fraction of missing information about : Both statistics and are helpful diagnostics for assessing how the missing data contribute to the uncertainty about . When the complete-data degrees of freedom are small, and there is only a modest proportion of missing data, the computed degrees of freedom, , can be much larger than , which is inappropriate. For example, with and , the computed degrees of freedom , which is inappropriate for data sets with complete-data degrees of freedom less than . Barnard and Rubin (1999) recommend the use of adjusted degrees of freedom where and . If you specify the complete-data degrees of freedom with the EDF= option, the MIANALYZE procedure uses the adjusted degrees of freedom, , for inference. Otherwise, the degrees of freedom are used.
Ist der Morbus Bechterew eine Folge der falschen Ernährung? Sicher nicht. Zwar kann eine lebenslang extrem falsche Ernährung das Immunsystem schon schwächen, so dass es insgesamt leichter Fehler machen könnte. Es gibt aber keinen nachweisbaren Zusammenhang zwischen dem Zustandekommen der Erkrankung und einer bestimmten Ernährungsweise bzw. einem bestimmten Nahrungsbestandteil. Wenn es wirklich so einfach wäre, dann hätte die medizinische Forschung das schon lange aufgedeckt. Menschen die Schweinefleisch meiden erkranken nicht seltener an Morbus Bechterew als Menschen die es essen. Muslime, welche aus religiösen Gründen auf Schweinefleisch verzichten, werden von der Krankheit nicht verschont. Eines muss man aber zugestehen: Vegetarier erkranken seltener an rheumatischen Erkrankungen überhaupt. Sicher spielt der Darm eine ganz entscheidende - und oft unterschätzte - Rolle für unser Immunsystem. Ist der MB erblich? Ist der MB eine Krankheit unserer Zeit? Ist der MB eine seltene Krankheit? In welchem Alter beginnt der MB? Kennt man die Ursache des MB? Soll man seine Nachkommen auf den Faktor HLA-B27 untersuchen lassen? Soll man als Morbus Bechterew-Betroffene/r überhaupt noch Kinder in die Welt setzen? Sponsoren Gasteiner Heilstollen Come back ins Leben Klinikum Bad Gastein Bewegungs Workshop Komm zu uns! Zeig Rückgrat! Indoor Zeig Rückgrat! Outdoor Socialmedia youtube75 45fcb © ÖVMB
Citrix Metaframe Access Issues TMAN2005 Registered I have been trying to access a corporate Citrix MetaFrame Server with no luck. I have tried using Safari, Firefox, and Internet Explorer. With Safari, I get an error with "Error opening ICA file" as the description in the "Client for Java Connection Center" window. There is no response from Safari and I have to force quit Safari. With Firefox, I get and error with the following description: SSL Error 0: You have not chosen to trust "/C=US/ST=/L=/O=Equifax/ OU=Equifax Secure Certificate Authority/CN=", the issuer of the server's security certificate. Error number: 183 I get the same error with Internet Explorer. I am running the Citrix ICA client for Mac OS X version 7.00.405 Any help would be greatly apprecitated. Thanks. peterthorn Registered TMAN2005 said: I finally figured it out. The solution: I found the appropriate certificate on www.geotrust.com and copied it into the keystore directory in the Citrix client directory structure. I have the exact same problem, but haven't been able to find the equifax certificate at geotrust. Could you be more specific on where to get it? Thanks! Peter falcon5182 Registered I'm also having issues with the Citrix ICA client. The problem I'm having is that the LAUNCH.ASP file tries to open with Photoshop or Dreamweaver. I can't get it open with the Citrix client. How can I change the file association so ASP files open the Citrix? Top
Category: How does a Solar Cell Work? Article Details • Written By: Michael Anissimov • Edited By: Niki Foster • Last Modified Date: 25 March 2016 • Copyright Protected: 2003-2016 Conjecture Corporation • Print this Article Free Widgets for your Site/Blog Subway is now the largest fast food chain in the world, with some 44,000 locations. more... April 30 , 1006 : The brightest supernova in recorded history appeared. more... There are a variety of different devices that use sunlight to generate power, but the basic way that a solar cell works is the same. In a photovoltaic (PV) cell, there are two layers of silicon, both of which are doped, or lightly mixed, with a certain element. Typically, one side is doped with boron and the other with arsenic. Because of the way each element bonds to the silicon, the layer containing boron, called the n-type layer, has a surplus of free electrons. The other side, the p-type layer, has a deficit of electrons, called holes. The p-type layer and n-type layer are pressed closely against each other and linked by a wire connected to an external load. This creates a circuit in the solar cell. When sunlight of the right energy level hits the n-type layer, which is on top, it excites some of the free electrons, which break loose from their natural state — pairs — and flow across the boundary between the layers to create a current. This only works if the two layers of the cell are pressed directly into each other. This is usually accomplished by fabricating both sides as part of the same process. Ad The current flows through the p-layer into the wire, which goes to the load, generally used to store electricity. Direct current (DC) is produced. If alternating current (AC) for household appliances is desired, the DC current is put through an alternator. After flowing through the load, the current continues back into the n-layer, which is lacking in electrons in some areas due to the current. The process continues. A current is generated without any mechanical input. Unfortunately, the materials used to make solar cells can be quite expensive. For protection, the top layer of the solar cell is covered with a glass plate affixed with transparent resin. The entire setup is called a p-n junction diode. More sophisticated cells use a series of p-n junction diodes. The first solar cells were only 1% efficient. Today, commercial solar panels are between 5% and 15% efficient. There are currently millions of dollars going into research on improving these percentages. Ad You might also Like Recommended Discuss this Article anon282231 Post 5 What is the role of a depletion zone in a solar cell? I mean, what happens to the depletion zone in the pn junction when photons strike it? wikesupply Post 4 I have been working on solar panel products for over two years, but before I read this article, I really didn't know how the solar cells generated electricity. PelesTears Post 3 @ Cougars- The silicon based solar cells that are so common today have only been around since the early fifties. These cells were produced initially for space missions to power satellites, space ships and the likes. Their value soon became evident, and they were soon adopted by the general public once the price started to come down. There is a new photovoltaic solar technology on the horizon, which may prove to be more efficient than flat silicon panels. Most home installs are not sun tracking, so they are most efficient when the sun is directly overhead, residential tracking systems can also be somewhat cost prohibitive, and often times HOAs and Covenants do not allow tracking systems. A few companies are working on systems that use Cadmium, Indium, Gallium, and diselenide to create panels comprised of tubes filled with photovoltaic materials. These systems can increase overall efficiency because they allow for peak absorption of solar radiation at multiple angles, extending the peak production times of the system. These systems look similar, but instead of individual cells, the panels look like they are holding a number of black, fluorescent light bulb like, tubes. cougars Post 2 How long has photovoltaic solar cell technology been available? Is photovoltaic solar technology a fairly new concept or has it been around for a while? I am in need of information on the history of solar power generation and I would appreciate it if someone could help me out. Thanks to anyone out there who knows. wikesupply Post 1 There are two sizes of solar cells: 125x125mm and 156x156mm. Each piece is rated power is 2.5W and 3.9W. For high quality solar cells, look for 10 years Peak Power Warranty - 90 percent minimum peak power or 20 years Peak Power Warranty - 80 percent minimum peak power. Post your comments Post Anonymously Login username password forgot password? Register username password confirm email
NCI Dictionary of Cancer Terms The NCI Dictionary of Cancer Terms features 8,442 terms related to cancer and medicine. We offer a widget that you can add to your website to let users look up cancer-related terms. Get NCI’s Dictionary of Cancer Terms Widget. Browse: humanized monoclonal antibody (HYOO-muh-nized MAH-noh-KLOH-nul AN-tee-BAH-dee) A type of antibody made in the laboratory by combining a human antibody with a small part of a mouse or rat monoclonal antibody. The mouse or rat part of the antibody binds to the target antigen, and the human part makes it less likely to be destroyed by the body's immune system.
Table of contents Retire an application 1. Archive database This is only relevant if the application has a database. You can skip this when retiring frontends. Grab a copy of the latest production backup of the database and put a copy in the /data/backups/archived directory on the backup-1.management machine. This will then be included in the sync to the offsite backup machines 2. Remove app from puppet Configure Puppet to remove the app from all servers. Change the app resource, and any database resources to ensure => absent, remove any host and load-balancer entries, but leave the hieradata entries. See this pull request for an example: After this has been deployed, remove it entirely. For example: 3. Remove smoke tests Remove any smokey tests specific to the application. 4. Remove deploy scripts Remove necessary scripts from the alphagov-deployment (private repo) and govuk-app-deployment repos. 5. Update Release app Mark the application as archived in the Release app. Edit the application in the release app (you’ll need the deploy permission to do this), and check the archived checkbox. This will hide it from the UI. 6. Remove from deploy Jenkins Remove entry from the deploy Jenkinses. This is managed through govuk-puppet in the common.yml. 7. Update Signon Mark the application as “retired” in signon, if it used it. Click on the Applications tab. Find the application that is being retired and click the “edit” button. Tick the box that says “This application is being retired”, then save your changes. 8. Update development scripts Remove from the development-vm directory Procfile and Pinfile: Leave a comment in the Procfile indicating that the port used to be used by this app, to avoid port conflicts causing a problem running this app locally in the future. 9. Check replication script Check the data replication scripts for anything specific to this application. Some applications have special case details in https://github.com/alphagov/env-sync-and-backup/. Any relating to the application should be removed. 10. Update DNS Request any public DNS entries be removed. If the app had an admin UI, it will have had public DNS entries in the publishing.service.gov.uk domain. Follow the instructions for DNS changes in order to remove these, and ask the Reliability Engineering team to approve any necessary Pull Requests. 11. Update docs Mark the application as retired in govuk-developer-docs. 12. Remove credentials Remove any hieradata credential entries for the app in govuk-secrets (private repo). 13. Drop database If Puppet hasn’t done it (eg for MongoDB databases), drop the database. 14. Remove jobs in CI If tests were set up, go to CI and choose “Delete Repository” for your project. 15. Remove other references Do a code search on GitHub to find any references to the application and update or remove them. 16. Unpublish routes Some applications are responsible for publishing certain routes. If you’re retiring a publishing application, make sure you check if any of its content items need to be unpublished and do it via the Publishing API. 17. Remove from Sentry Since the application has been retired, it shouldn’t be tracked in Sentry. 18. Archive the repo Go into the repository settings in Github, and archive the repo. This page was last reviewed . It needs to be reviewed again by the page owner #govuk-2ndline.
Question about 1993 Lexus Ls 400 1 Answer Side bulb blown How do I change the bulb on the front side light? I tried to take the headlamp out but it seems like a major dismantling job, surely there is an easier way. I took it to Halfords and they didn't know. Posted by on 1 Answer • Level 3: An expert who has achieved level 3 by getting 1000 points All-Star: An expert that got 10 achievements. MVP: An expert that got 5 achievements. Vice President: An expert whose answer got voted for 100 times. • Master • 521 Answers Ok, open the hood, just at upper corner the side light have an small phillip bolt ,loose this and introduce your finger between headlamp and side lamp, then pull to front with care but firm. for change bulbs in bumper , remove cover under the engine and then have acces to lamps Posted on Apr 22, 2009 1 Suggested Answer 6ya6ya • 2 Answers SOURCE: I have freestanding Series 8 dishwasher. Lately during the filling cycle water hammer is occurring. How can this be resolved Hi, a 6ya expert can help you resolve that issue over the phone in a minute or two. best thing about this new service is that you are never placed on hold and get to talk to real repairmen in the US. the service is completely free and covers almost anything you can think of (from cars to computers, handyman, and even drones). click here to download the app (for users in the US for now) and get all the help you need. goodluck! Posted on Jan 02, 2017 Add Your Answer Uploading: 0% my-video-file.mp4 Complete. Click "Add" to insert your video. Add × Loading... Loading... Related Questions: 1 Answer How to replace drivers side front headlight unit on 2003 vauxhaull vectra The bulbs are little devils to get hold off due to the lack of space and light inside the bonnet. So when dealing with the main beam pull out the side bulb and turn on the side lights. This will give you some light in the tight space. Before you undo the spring clip, attach a piece of fairly rigid plastic tubing (about 6 inches long) to the bulbs terminal. Make sure that the tubing is a firm fit, then undo the clip and withdraw the bulb using the plastic tube. This should insure that you don't drop the bulb into the headlamp cover or the car's floor pan like I did. Also notice which way around the bulb is located i.e.: the bulb flange is "D" shaped and can only be fitted one way round. Use the plastic tube to grip the terminal of the new bulb and guide it into the bulb's headlamp slot. Make sure that the flange is located correctly with your fingers, and then apply firm pressure to the tube to hold the bulb in place and relocate the spring clip one side at a time. The covers on the back of my headlights were just made of rubber and just pulled off rather than screwed. The job is awkward and more touchy feely than seeing what you are doing, but it two bulbs can be changed in under an hour Jul 11, 2012 \| Vauxhall Vectra Cars & Trucks 1 Answer How to replace bulb in front bumper turn signal housing 1995 jag xj6 hello, • Identify the blown bulb. Note that if a headlight bulb has blown, the dip beam is on the outside and the high beam is on the inside. Turn off the ignition switch and light switches. • Open the hood and replace any blown headlight bulbs. Press the plastic spring clips and remove the bulb holder from the assembly. Pull the bulb out of the bulb holder and replace it with a new bulb. It only fits one way. Be sure not to touch the bulb's glass. Replace any parking lights at the same time, they are in the headlamp unit too. • Remove the front turn signal light unit in the bumper to change the turn signal bulb. Push gently on the inside edge and insert a blade between the bumper and the lens at the outside edge. Compress the spring and remove the lens. Fit a new lamp by inserting it into the plug. The side marker light is changed using a similar process. • Open the trunk to replace the rear fog light, stop light, tail light, reversing light and turn signal light. Remove the rear light cover and turn the bulb holder fastener 90 degrees counter clockwise. Remove the expired bulb, fit a new one and replace the parts, turning the bulb holder clockwise. • Thanks.. May 21, 2011 \| 1995 Jaguar XJ6 1 Answer 2005 chevy equinox headlights went out, only the driver side bright light works. Does this mean the passenger side headlamp is blown? Sounds like you are right own...to change each bulb...you need to remove the chrome grill from the front center...be sure to pry up the clips carefully so not to break the trim bar...then each headlight assembly is removable with 3 10 mm bolts...do not touch your exposed fingers on the new bulbs...yes the low/high bulb is one unit...that is why I believe it is just your bulbs...about $11. each...Hope this helps. Jan 25, 2011 \| 2005 Chevrolet Equinox 1 Answer Headlights wont come on. Running and tail lights fine. Wont come on even if manually put them on. Emerg brake not on, fuse seems fine... Is neither the high beam nor low beam working? Neither the left side nor the right side for each? Knowing this could help isolate the problem. It could be anything from a simple case of blown headlamp bulbs to a burned-up headlamp switch to a defective Daytime Running Lamp Module. I will say, however that I have seen quite a few problems with the headlamp bulb sockets on several of the GMs. Jan 22, 2011 \| 2005 Buick Century 1 Answer 1997 ford contour tail lights won't work but brake lights do already changed bulbs what fuse is for parking light There are 2 fuses,left and right, so I dont think both could be out? fuses. 33 and 35. make sure they have power on both sides of the fuse. If one side has power, its blown, if no power, the headlamp switch is bad, or bad connection. The RED/BLACK wires coming out of the headlamp switch go to those fuses. 33-35 Dec 10, 2010 \| 1997 Ford Contour 1 Answer Changing Low Beam Headlamp Replacement 2005 Mazda 3. Check out the following: It's not easy. You need a flashlight. First, take the batter vent assembly off by removing the screw over the headlamp assembly. Open the battery box by snapping off the tabs, lift out, and move the vent to the side. Second, disconnect the headlight wire by pressing down on the opposing tabs located on the side of the connector and pull straight back. Move the wire to the side. Next, peel the rubber fitting around the bulb (located on the back of the headlamp assembly). Look inside the headlamp assembly with a flashlight to see what you got yourself into. You'll have to unhook the retainer wire that holds the bulb in place. This requires you to push the top part of the wire toward the front of the car and lift up. The wire should swing to the left (passenger side) releasing the bulb and bulb housing. Pull the old bulb out of the housing (it might be tight), replace with the new one (try not to touch the glass). Replace the bulb/housing. Make sure the little tap fits back into its slot (upper diver side). Push the wire clamp back over the bulb, pushing up back and down making sure it's hooked and your done. It seems easy once you've done it. Good luck! EJ Kudelka Nov 09, 2010 \| 2005 Mazda 3 1 Answer Need help changing out the low beam on drivers side of 2006 buick rainier Removal & Installation Bulb Replacement To Remove: CAUTION Halogen bulbs contain pressurized gas. Mishandling a bulb may result in the bulb shattering into flying glass fragments. CAUTION Make sure to wear eye protection when handling a halogen bulb. CAUTION Do not touch the glass of a halogen bulb, only handle the bulb by the base. CAUTION Do not allow a halogen bulb to come into contact with dirt or moisture. CAUTION Make sure to dispose of a used halogen bulb properly. CAUTION Do not allow children to handle halogen bulbs. 1. Turn the lamp switch to the off position. 2. Make sure the bulb is cool before changing. 3. Remove the headlamp assembly from the headlamp panel. 4. Remove the rear access cover from the headlamp assembly. 5. Disconnect the headlamp connector. 6. Remove the bulb from the headlamp assembly by rotating the bulb and socket counterclockwise. To Install: 1. Connect the headlamp connector. 2. Install the bulb into the headlamp assembly by rotating the bulb and socket clockwise. 3. Install the rear access cover onto the headlamp assembly. 4. Install the headlamp assembly into the headlamp panel. Headlamp Assembly Chevrolet To Remove: Headlamp Assembly (Chevrolet) gm-06-00-1591.gif 1. Remove the grille from the front of the vehicle. 2. Release the headlamp assembly retaining tabs. 3. If equipped, disconnect the headlamp leveling motor connector. 4. Disconnect the headlamp connector. 5. Remove the park/turn and side marker bulb sockets from the headlamp assembly. 6. Remove the headlamp assembly from the headlamp panel. To Install: 1. Place the headlamp assembly into the headlamp panel. 2. Install the park/turn and side marker bulb sockets into the headlamp assembly. 3. Connect the headlamp connector. 4. If equipped, connect the headlamp leveling motor connector. 5. Lock the headlamp assembly retaining tabs. 6. Install the grille onto the front of the vehicle. GMC, Oldsmobile, Buick, Isuzu, and Saab To Remove: Headlamp Assembly (GMC, Saab, Oldsmobile, and Buick) gm-06-00-1590.gif 1. Release the headlamp assembly retaining tabs. 2. Disconnect the headlamp connector. 3. Remove the park/turn bulb socket from the headlamp assembly. 4. For GMC, Saab, Oldsmobile, and Isuzu vehicles, remove the side marker bulb socket from the headlamp assembly. 5. Remove the headlamp assembly from the headlamp panel. To Install: 1. Place the headlamp assembly into the headlamp panel. 2. For GMC, Saab, Oldsmobile, and Isuzu vehicles, install the side marker bulb socket into the headlamp assembly. 3. Install the park/turn bulb socket into the headlamp assembly. 4. Connect the headlamp connector. 5. Lock the headlamp assembly retaining tabs. prev.gif next.gif Oct 28, 2010 \| 2006 Buick Rainier 1 Answer How to replace headlight bulb on 2002 chevy trailblazer Headlamp Removal & Installation Bulb Replacement To Remove: CAUTION Halogen bulbs contain pressurized gas. Mishandling a bulb may result in the bulb shattering into flying glass fragments. CAUTION Make sure to wear eye protection when handling a halogen bulb. CAUTION Do not touch the glass of a halogen bulb, only handle the bulb by the base. CAUTION Do not allow a halogen bulb to come into contact with dirt or moisture. CAUTION Make sure to dispose of a used halogen bulb properly. CAUTION Do not allow children to handle halogen bulbs. 1. Turn the lamp switch to the off position. 2. Make sure the bulb is cool before changing. 3. Remove the headlamp assembly from the headlamp panel. 4. Remove the rear access cover from the headlamp assembly. 5. Disconnect the headlamp connector. 6. Remove the bulb from the headlamp assembly by rotating the bulb and socket counterclockwise. To Install: 1. Connect the headlamp connector. 2. Install the bulb into the headlamp assembly by rotating the bulb and socket clockwise. 3. Install the rear access cover onto the headlamp assembly. 4. Install the headlamp assembly into the headlamp panel. Headlamp Assembly Chevrolet To Remove: Headlamp Assembly (Chevrolet) gm-06-00-1591.gif 1. Remove the grille from the front of the vehicle. 2. Release the headlamp assembly retaining tabs. 3. If equipped, disconnect the headlamp leveling motor connector. 4. Disconnect the headlamp connector. 5. Remove the park/turn and side marker bulb sockets from the headlamp assembly. 6. Remove the headlamp assembly from the headlamp panel. To Install: 1. Place the headlamp assembly into the headlamp panel. 2. Install the park/turn and side marker bulb sockets into the headlamp assembly. 3. Connect the headlamp connector. 4. If equipped, connect the headlamp leveling motor connector. 5. Lock the headlamp assembly retaining tabs. 6. Install the grille onto the front of the vehicle. Jul 20, 2010 \| 2002 Chevrolet TrailBlazer 1 Answer How to replace front turn light bulb on Caddy SRX 2004? 1. Open the hood. 2. Remove the push-pin retainers that secure the top of the upper fascia and grille assembly to the upper tie bar to allow the upper fascia and grille assembly to flex forward. 3. Remove the bolt and the push-pin retainer that secures the top of the headlamp assembly to the upper tie bar. 1. Gently pull the upper fascia and grille assembly forward only far enough to remove the bolt located at the bottom inboard side of the headlamp assembly. 2. Pull the headlamp assembly straight out to disengage the snap-in retainer at the lower outboard back surface of the assembly. Turn the assembly so that the front inboard corner clears the front fascia. 3. Pull the headlamp assembly out and away from the vehicle 4. Disconnect the headlamp electrical connector. 5. Remove the headlamp assembly from the vehicle. 6. Remove the headlamp bulb cover from the headlamp assembly. 7. Remove the headlamp bulb socket from the headlamp assembly. 8. Remove the headlamp bulb. 9. Remove the side marker bulb socket from the outer side of the headlamp assembly. 10. Remove the side marker bulb. Feb 12, 2009 \| 2006 Cadillac SRX 2 Answers Changing parking light bulb in front Fisrt of all remove the Plastic fixings from the top of grill then carefully place your hands in the bottom grill slots and give a sharp pull,this should release the inner push type fixings (4 in all, one on either side of the headlamp towards middle of car and one either side at the wing)(very tight fit) the to release the outer edge ones carefully run your hand along the bottom of the headlamp and behind the plastic bottom edge of the grill the till you get as close to the wing as possible,then another sharp pull.Do this again for the other side and you good to go.On refiting make sure the two wing end pins are placed in first as these can be a pain if not inserted properly.. Hope this helps Sep 26, 2008 \| 2004 Suzuki Grand Vitara Not finding what you are looking for? Cars & Trucks Logo 228 people viewed this question Ask a Question Usually answered in minutes! Top Lexus Experts yadayada yadayada Level 3 Expert 69199 Answers ROBERT GARCIA ROBERT GARCIA Level 3 Expert 521 Answers Colin Stickland Colin Stickland Level 3 Expert 22026 Answers Are you a Lexus Expert? Answer questions, earn points and help others Answer questions Manuals & User Guides Loading...
Bone Biopsy What is a Bone Biopsy? A bone biopsy is an image-guided procedure in which a small sample of a bone is removed from the body and examined under a microscope. The procedure is often called a closed or needle bone biopsy, because it involves inserting a needle directly into the bone. Computed tomography (CT) or, in some cases, magnetic resonance imaging (MRI) is used to guide the procedure. What are some common uses of the procedure? View larger with caption Computed Tomography (CT) image Bone biopsies are performed to: • confirm the diagnosis of a bone disorder • investigate an abnormal area, or lesion, seen on x-ray, bone scan, CT or MRI • distinguish bone tumor from other conditions, such an infection • distinguish whether a tumor is benign or cancerous • determine the cause of an infection or inflammation • identify the cause of bone pain. How should I prepare? Prior to your procedure, your blood may be tested to determine how well your kidneys are functioning and whether your blood clots normally. You may be instructed not eat or drink for eight hours before your biopsy. However, you may take your routine medications with sips of water. If you are diabetic and take insulin, you should talk to your doctor as your usual insulin dose may need to be adjusted. Prior to a bone biopsy, you should report to your doctor all medications that you are taking, including herbal supplements, and if you have any allergies, especially to anesthesia. Your physician may advise you to stop taking aspirin or a blood thinner three days before your procedure. Also, inform your doctor about recent illnesses and other medical conditions. You may be asked to wear a gown during the procedure. Women should always inform their physician if there is any possibility that they are pregnant. Some procedures using image-guidance are typically not performed during pregnancy because radiation can be harmful to the fetus. See the Safety Page (www.RadiologyInfo.org/safety/) for more information on pregnancy and x-rays. You may be asked to remove some or all of your clothes and to wear a gown during the exam. You may also be asked to remove jewelry, eye glasses and any metal objects or clothing that might interfere with the x-ray images. You may want to have a relative or friend accompany you and drive you home afterward. This is necessary if you have been sedated. Preparation for a biopsy procedure will be similar for children. If your child is undergoing a biopsy procedure, the physician will provide you with instructions. What does the equipment look like? View larger with caption Photograph of needles and drill tips used during a bone biopsy A special drill needle is used for a closed bone biopsy. This needle is generally several inches long with a hollow core to capture the bone specimen. The CT scanner is typically a large, box-like machine with a hole, or short tunnel, in the center. You will lie on a narrow examination table that slides into and out of this tunnel. Rotating around you, the x-ray tube and electronic x-ray detectors are located opposite each other in a ring, called a gantry. The computer workstation that processes the imaging information is located in a separate control room, where the technologist operates the scanner and monitors your examination in direct visual contact and usually with the ability to hear and talk to you with the use of a speaker and microphone. Other equipment that may be used during the procedure includes an intravenous line (IV), ultrasound machine and devices that monitor your heart beat and blood pressure. How does the procedure work? The physician inserts a needle through the skin and advances it into the bone. A second needle, inserted through the first needle, removes a sample of the bone. The needles are then removed. The procedure is usually image-guided. How is the procedure performed? Bone biopsies are usually done on an outpatient basis. You will be positioned so that the physician can easily reach the bone that is to be sampled. A belt or strap may be used to hold you in the correct position. If the procedure is performed with CT, you will lie down during the procedure. A limited CT scan will be performed to confirm the location to be biopsied You may be connected to monitors that track your heart rate, blood pressure and pulse during the procedure. A nurse or technologist may insert an intravenous (IV) line into a vein in your hand or arm so that sedation or relaxation medication may be given intravenously during the procedure. You may be also given a mild sedative prior to the biopsy. A local anesthesia will be injected to numb the path of the needle. A very small nick is made in the skin at the site where the biopsy needle is to be inserted. View larger with caption Illustration showing the route used for a bone puncture. Using image-guidance, the physician will insert the needle through the skin, advance it to the bone and then insert a second needle through the first needle, which will remove a small sample of the lesion into its hollow core. As the needle being advanced towards the lesion, additional limited CT images may be obtained to monitor the passage of the needle. After the sampling, the needle will be removed. Pressure will be applied to prevent any bleeding and the opening in the skin is covered with a bandage. No sutures are needed. Your intravenous line will be removed. You may be taken to an observation area for several hours. X-ray(s) or other imaging tests may be performed to monitor for complications. A needle biopsy is usually completed within 30 to 60 minutes but may take longer, depending on the size of the biopsied lesion and on the difficulty reaching it with the needle. What will I experience during and after the procedure? When you receive the local anesthetic to numb the skin, you will feel a slight pin prick from the needle. You may feel some pressure when the biopsy needle is inserted and aching pain or pressure when the bone sample is removed. After the procedure, the biopsy site may be sore for up to a week. You should talk to your doctor about pain medication. You should call your doctor if there is excessive bleeding from the biopsy site, or signs of infection such as: • increased pain, swelling, redness or warmth • pus draining from the site • swollen lymph nodes in the neck, armpit or groin • fever or chills. Who interprets the results and how do I get them? The specimen obtained by the radiologist will be analyzed by pathologists and/or microbiologists, and it may take up to five to seven days to get the final result. The ordering physician will communicate the results to you. What are the benefits vs. risks? Benefits • Needle biopsy is a reliable method of obtaining tissue samples that can help diagnose whether a lesion is benign (non-cancerous) or malignant. • A closed needle biopsy is less invasive than surgical biopsy and can be performed using local anesthesia and moderate (conscious) sedation, while the surgical biopsy involves a larger incision in the skin and usually requires general anesthesia. • Generally, the procedure is not painful and the results are as accurate as when a bone sample is removed surgically. • Recovery time is brief. Risks • Any procedure where the skin is penetrated carries a risk of infection. The chance of infection requiring antibiotic treatment appears to be less than one in 1,000. • Complications following a bone biopsy are rare. However, there is a small chance the biopsy needle may break the bone or injure a nerve, blood vessel or organ nearby. There is a very small chance that the bone may become infected or weak and not heal properly. What are the limitations of Bone Biopsy? A bone biopsy may not be able to be performed on patients who: • are unable to lie still during the procedure. • have a condition affecting the immune system, which increases the chances of an infection at the biopsy site. • take aspirin or blood thinners or have a bleeding disorder, which may increase chances for bleeding at the biopsy site. It may be difficult to remove an adequate sample of bone tissue with a needle biopsy. Did you find the information you were looking for? If you wish to submit a comment, click here. Locate an ACR-accredited provider: To locate a medical imaging or radiation oncology provider in your community, you can search the ACR-accredited facilities database. Interventional radiology: For more information on interventional radiology procedures, visit the Society of Interventional Radiology (SIR) website at www.sirweb.org. This website does not provide costs for exams. The costs for specific medical imaging tests and treatments vary widely across geographic regions. Many—but not all—imaging procedures are covered by insurance. Discuss the fees associated with your medical imaging procedure with your doctor and/or the medical facility staff to get a better understanding of the portions covered by insurance and the possible charges that you will incur. Web page review process: This Web page is reviewed regularly by a physician with expertise in the medical area presented and is further reviewed by committees from the American College of Radiology (ACR) and the Radiological Society of North America (RSNA), comprising physicians with expertise in several radiologic areas. Outside links: For the convenience of our users, RadiologyInfo.org provides links to relevant websites. RadiologyInfo.org, ACR and RSNA are not responsible for the content contained on the web pages found at these links. Images: Images are shown for illustrative purposes. Do not attempt to draw conclusions or make diagnoses by comparing these images to other medical images, particularly your own. Only qualified physicians should interpret images; the radiologist is the physician expert trained in medical imaging. 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<< >> Ãëàâà 1 ÕÈÌÈß ÁÅËÊÎÂ Ìèð ñàìîãî ñëîæíîãî—æèçíü. Í.Í. Ñåìåíîâ Æèâîé îðãàíèçì õàðàêòåðèçóåòñÿ âûñøåé ñòåïåíüþ óïîðÿäî÷åííîñòè ñîñòàâëÿþùèõ åãî èíãðåäèåíòîâ è óíèêàëüíîé ñòðóêòóðíîé îðãàíèçàöèåé, îáåñïå÷èâàþùåé êàê åãî ôåíîòèïè÷åñêèå ïðèçíàêè, òàê è ìíîãîîáðàçèå áèîëîãè÷åñêèõ ôóíêöèé. Â ýòîì ñòðóêòóðíî-ôóíêöèîíàëüíîì åäèíñòâå îðãàíèçìîâ, ñîñòàâëÿþùåì ñóùíîñòü æèçíè, áåëêè (áåëêîâûå òåëà) èãðàþò âàæíåéøóþ ðîëü, íå çàìåíÿåìóþ äðóãèìè îðãàíè÷åñêèìè ñîåäè-íåíèÿìè. Áåëêè—ýòî âûñîêîìîëåêóëÿðíûå àçîòñîäåðæàùèå îðãàíè÷åñêèå âåùåñòâà, ìîëåêóëû êîòîðûõ ïîñòðîåíû èç îñòàòêîâ àìèíîêèñëîò. Íàçâàíèå «ïðîòåèíû» (îò ãðå÷. ðãî1î§—ïåðâûé, âàæíåéøèé), ïî-âèäèìîìó, áîëåå òî÷íî îòðàæàåò ïåðâîñòåïåííîå áèîëîãè÷åñêîå çíà÷åíèå ýòîãî êëàññà âåùåñòâ. Ïðèíÿòûå â îòå÷åñòâåííîé ëèòåðàòóðå òåðìèíû «áåëêè» è «áåëêîâûå âåùåñòâà» ñâÿçàíû ñ îáíàðóæåíèåì â òêàíÿõ æèâîòíûõ è ðàñòåíèé âåùåñòâ, èìåþùèõ ñõîäñòâî ñ áåëêîì êóðèíîãî ÿéöà. Â íàøå âðåìÿ, êîãäà àáñîëþòíî äîñòîâåðíî óñòàíîâëåíî, ÷òî íàñëåäñòâåííàÿ èíôîðìàöèÿ ñîñðåäîòî÷åíà â ìîëåêóëå ÄÍÊ êëåòîê ëþáûõ æèâûõ îðãàíèçìîâ, íå âûçûâàåò ñîìíåíèÿ, ÷òî òîëüêî áåëêè ÿâëÿþòñÿ òåìè ìîëåêóëÿðíûìè èíñòðóìåíòàìè, ïðè ïîìîùè êîòîðûõ ðåàëèçóåòñÿ ãåíåòè÷åñêàÿ èíôîðìàöèÿ. Áåç áåëêîâ, â ÷àñòíîñòè ôåðìåíòîâ, ÄÍÊ íå ìîæåò ðåïëèöèðîâàòüñÿ, íå ìîæåò ñàìîâîñïðî- èçâîäèòüñÿ, ò.å. ëèøåíà ñïîñîáíîñòè ïåðåäàâàòü ãåíåòè÷åñêóþ èíôîðìàöèþ. Æèâàÿ ïðèðîäà õàðàêòåðèçóåòñÿ ðÿäîì ñâîéñòâ, îòëè÷àþùèõ åå îò íåæèâîé ïðèðîäû, è ïî÷òè âñå ýòè ñâîéñòâà ñâÿçàíû ñ áåëêàìè. Ïðåæäå âñåãî äëÿ æèâûõ îðãàíèçìîâ õàðàêòåðíû øèðîêîå ðàçíîîáðàçèå áåëêîâûõ ñòðóêòóð è èõ âûñîêàÿ óïîðÿäî÷åííîñòü; ïîñëåäíÿÿ ñóùåñòâóåò âî âðåìåíè è ïðîñòðàíñòâå. Óäèâèòåëüíàÿ ñïîñîáíîñòü æèâûõ îðãàíèçìîâ ê âîñïðîèçâåäåíèþ ñåáå ïîäîáíûõ òàêæå ñâÿçàíà ñ áåëêàìè. Ñîêðàòèìîñòü, äâèæåíèå — íåïðåìåííûå àòðèáóòû æèâûõ ñèñòåì—èìåþò ïðÿìîå îòíîøåíèå ê áåëêîâûì ñòðóêòóðàì ìûøå÷íîãî àïïàðàòà. Íàêîíåö, æèçíü íåìûñëèìà áåç îáìåíà âåùåñòâ, ïîñòîÿííîãî îáíîâëåíèÿ ñîñòàâíûõ ÷àñòåé æèâîãî îðãà-íèçìà, ò.å. áåç ïðîöåññîâ àíàáîëèçìà è êàòàáîëèçìà (ýòîãî óäèâèòåëüíîãî åäèíñòâà ïðîòèâîïîëîæíîñòåé æèâîãî), â îñíîâå êîòîðûõ ëåæèò äåÿòåëüíîñòü êàòàëèòè÷åñêè àêòèâíûõ áåëêîâ—ôåðìåíòîâ. Òàêèì îáðàçîì, áåëêè (áåëêîâûå âåùåñòâà) ñîñòàâëÿþò îñíîâó è ñòðóêòóðû, è ôóíêöèè æèâûõ îðãàíèçìîâ. Ïî îáðàçíîìó âûðàæåíèþ îäíîãî èç îñíîâîïîëîæíèêîâ ìîëåêóëÿðíîé áèîëîãèè Ô. Êðèêà, áåëêè âàæíû ïðåæäå âñåãî ïîòîìó, ÷òî îíè ìîãóò âûïîëíÿòü ñàìûå ðàçíîîáðàçíûå ôóíêöèè, ïðè÷åì ñ íåîáûêíîâåííîé ëåãêîñòüþ è èçÿùåñòâîì. Ïîäñ÷èòàíî, ÷òî â ïðèðîäå ïðèìåðíî 1010—1012 ðàçëè÷íûõ áåëêîâ, îáåñïå÷èâàþùèõ ñóùåñòâîâàíèå îêîëî 106 âèäîâ æèâûõ îðãàíèçìîâ ðàçëè÷íîé ñëîæíîñòè îðãà-íèçàöèè íà÷èíàÿ îò âèðóñîâ è êîí÷àÿ ÷åëîâåêîì. Èç ýòîãî îãðîìíîãî êîëè÷åñòâà ïðèðîäíûõ áåëêîâ èçâåñòíû òî÷íîå ñòðîåíèå è ñòðóêòóðà íè÷òîæíî ìàëîé ÷àñòè (ñì. äàëåå). Êàæäûé îðãàíèçì õàðàêòåðèçóåòñÿ óíèêàëüíûì íàáîðîì áåëêîâ. Ôåíîòèïè÷åñêèå ïðèçíàêè è ìíîãîîáðàçèå ôóíêöèé îáóñëîâëåíû ñïåöèôè÷íîñòüþ îáúåäèíåíèÿ ýòèõ áåëêîâ, âî ìíîãèõ ñëó÷àÿõ â âèäå íàä- è ìóëüòèìîëåêóëÿðíûõ ñòðóêòóð, â ñâîþ î÷åðåäü îïðåäåëÿþùèõ óëüòðàñòðóêòóðó êëåòîê è èõ îðãàíåëë. Â êëåòêå Å.ñîÍ ñîäåðæèòñÿ îêîëî 3000 ðàçëè÷íûõ áåëêîâ, à â îðãàíèçìå ÷åëîâåêà íàñ÷èòûâàåòñÿ áîëåå 100000 ðàçíîîáðàçíûõ áåëêîâ. Ñàìîå óäèâèòåëüíîå, ÷òî âñå ïðèðîäíûå áåëêè ñîñòîÿò èç íåáîëüøîãî ÷èñëà ñðàâíèòåëüíî ïðîñòûõ ñòðóêòóðíûõ áëîêîâ, ïðåäñòàâëåííûõ ìîíîìåðíûìè ìîëåêóëàìè —àìèíîêèñëîòàìè, ñâÿçàííûìè äðóã ñ äðóãîì â ïîëèïåïòèäíûå öåïè. Ïðèðîäíûå áåëêè ïîñòðîåíû èç 20 ðàçëè÷íûõ àìèíîêèñëîò. Ýòè àìèíîêèñëîòû ìîãóò îáúåäèíÿòüñÿ â ñàìîé ðàçíîé ïîñëåäîâàòåëüíîñòè, ïîýòîìó îíè ìîãóò îáðàçîâûâàòü ãðîìàäíîå êîëè÷åñòâî ðàçíîîáðàçíûõ áåëêîâ. ×èñëî èçîìåðîâ, êîòîðîå ìîæíî ïîëó÷èòü ïðè âñåâîçìîæíûõ ïåðåñòàíîâêàõ óêàçàííîãî ÷èñëà àìèíîêèñëîò â ïîëèïåïòèäå, èñ÷èñëÿåòñÿ îãðîìíûìè âåëè÷èíàìè. Òàê, åñëè èç 2 àìèíîêèñëîò âîçìîæíî îáðàçîâàíèå òîëüêî äâóõ èçîìåðîâ, òî óæå èç 4 àìèíîêèñëîò òåîðåòè1÷8 åñêè âîçìîæíî îáðàçîâàíèå 24 èçîìåðîâ, à èç 20 àìèíîêèñëîò — 2,4 • 1018 ðàçíîîáðàçíûõ áåëêîâ. Íåòðóäíî ïðåäâèäåòü, ÷òî ïðè óâåëè÷åíèè ÷èñëà ïîâòîðÿþùèõñÿ àìèíîêèñëîòíûõ îñòàòêîâ â áåëêîâîé ìîëåêóëå ÷èñëî âîçìîæíûõ èçîìåðîâ âîçðàñòàåò äî àñòðîíîìè÷åñêèõ âåëè÷èí. ßñíî, ÷òî ïðèðîäà íå ìîæåò ïîçâîëèòü ñëó÷àéíûõ ñî÷åòàíèé àìèíîêèñëîòíûõ ïîñëåäîâàòåëüíîñòåé è äëÿ êàæäîãî âèäà õàðàêòåðåí ñâîé ñïåöèôè÷åñêèé íàáîð áåëêîâ, îïðåäåëÿåìûé, êàê òåïåðü èçâåñòíî, íàñëåäñòâåííîé èíôîðìàöèåé, çàêîäèðîâàííîé â ìîëåêóëå ÄÍÊ æèâûõ îðãàíèçìîâ. Èìåííî èíôîðìàöèÿ, ñîäåðæàùàÿñÿ â ëèíåéíîé ïîñëåäîâàòåëüíîñòè íóêëåîòèäîâ ÄÍÊ, îïðåäåëÿåò ëèíåéíóþ ïîñëåäîâàòåëüíîñòü îñòàòêîâ àìèíîêèñëîò â ïîëèïåïòèäíîé öåïè ñèíòåçè-ðóåìîãî áåëêà. Îáðàçîâàâøàÿñÿ ëèíåéíàÿ ïîëèïåïòèäíàÿ öåïü ñàìà òåïåðü îêàçûâàåòñÿ íàäåëåííîé ôóíêöèîíàëüíîé èíôîðìàöèåé, â ñîîòâåòñòâèè ñ êîòîðîé îíà ñàìîïðîèçâîëüíî ïðåîáðàçóåòñÿ â îïðåäåëåííóþ ñòàáèëüíóþ òðåõìåðíóþ ñòðóêòóðó. Òàêèì îáðàçîì, ëàáèëüíàÿ ïîëèïåïòèäíàÿ öåïü ñêëàäûâàåòñÿ, ñêðó÷èâàåòñÿ â ïðîñòðàíñòâåííóþ ñòðóêòóðó áåëêîâîé ìîëå-êóëû, ïðè÷åì íå õàîòè÷íî, à â ñòðîãîì ñîîòâåòñòâèè ñ èíôîðìàöèåé, ñîäåðæàùåéñÿ â ïîñëåäîâàòåëüíîñòè àìèíîêèñëîòíûõ îñòàòêîâ. Ó÷èòûâàÿ âåäóùóþ ðîëü áåëêîâ â æèâîé ïðèðîäå è òîò ôàêò, ÷òî áåëêè, ñîñòàâëÿÿ ïî÷òè ïîëîâèíó ñóõîé ìàññû æèâîãî îðãàíèçìà, íàäåëåíû óäèâèòåëüíûì ðàçíîîáðàçèåì ôóíêöèé, èçó÷åíèå êóðñà áèîõèìèè â ìåäèöèíñêèõ âûñøèõ ó÷åáíûõ çàâåäåíèÿõ îáû÷íî íà÷èíàþò ñ ýòîãî êëàññà îðãàíè÷åñêèõ âåùåñòâ. ÔÓÍÊÖÈÈ ÁÅËÊÎÂ Áåëêè âûïîëíÿþò ìíîæåñòâî ñàìûõ ðàçíîîáðàçíûõ ôóíêöèé, õàðàêòåðíûõ äëÿ æèâûõ îðãàíèçìîâ, ñ íåêîòîðûìè èç êîòîðûõ ìû ïîçíàêîìèìñÿ áîëåå ïîäðîáíî ïðè äàëüíåéøåì èçó÷åíèè êóðñà. Íèæå ðàññìàòðèâàþòñÿ ãëàâíûå è â íåêîòîðîì ñìûñëå óíèêàëüíûå áèîëîãè÷åñêèå ôóíêöèè áåëêîâ, íåñâîéñòâåííûå èëè ëèøü ÷àñòè÷íî ïðèñóùèå äðóãèì êëàññàì áèîïîëèìåðîâ. Êàòàëèòè÷åñêàÿ ôóíêöèÿ. Ê 1995 ã. áûëî èäåíòèôèöèðîâàíî áîëåå 3400 ôåðìåíòîâ. Áîëüøèíñòâî èçâåñòíûõ â íàñòîÿùåå âðåìÿ ôåðìåíòîâ, íàçûâàåìûõ áèîëîãè÷åñêèìè êàòàëèçàòîðàìè, ÿâëÿåòñÿ áåëêàìè. Ýòà ôóíêöèÿ áåëêîâ, õîòÿ è íå îêàçàëàñü óíèêàëüíîé, îïðåäåëÿåò ñêîðîñòü õèìè÷åñêèõ ðåàêöèé â áèîëîãè÷åñêèõ ñèñòåìàõ . Òðàíñïîðòíàÿ ôóíêöèÿ. Äûõàòåëüíàÿ ôóíêöèÿ êðîâè, â ÷àñòíîñòè ïåðåíîñ êèñëîðîäà, îñóùåñòâëÿåòñÿ ìîëåêóëàìè ãåìîãëîáèíà—áåëêà ýðèòðîöèòîâ. Â òðàíñïîðòå ëèïèäîâ ïðèíèìàþò ó÷àñòèå àëüáóìèíû ñûâîðîòêè êðîâè. Ðÿä äðóãèõ ñûâîðîòî÷íûõ áåëêîâ îáðàçóåò êîìïëåêñû ñ æèðàìè, ìåäüþ, æåëåçîì, òèðîêñèíîì, âèòàìèíîì À è äðóãèìè ñîåäèíåíèÿìè, îáåñïå÷èâàÿ èõ äîñòàâêó â ñîîòâåòñòâóþùèå îðãàíû-ìèøåíè. Çàùèòíàÿ ôóíêöèÿ. Îñíîâíóþ ôóíêöèþ çàùèòû â îðãàíèçìå âûïîëíÿåò èììóííàÿ ñèñòåìà, êîòîðàÿ îáåñïå÷èâàåò ñèíòåç ñïåöèôè÷åñêèõ çàùèòíûõ áåëêîâ-àíòèòåë â îòâåò íà ïîñòóïëåíèå â îðãàíèçì áàêòåðèé, òîêñèíîâ, âèðóñîâ èëè ÷óæåðîäíûõ áåëêîâ. Âûñîêàÿ ñïåöèôè÷íîñòü âçàèìîäåéñòâèÿ àíòèòåë ñ àíòèãåíàìè (÷óæåðîäíûìè âåùåñòâàìè) ïî òèïó áåëîê-áåëêîâîå âçàèìîäåéñòâèå ñïîñîáñòâóåò óçíàâàíèþ è íåéòðàëèçàöèè áèîëîãè÷åñêîãî äåéñòâèÿ àíòèãåíîâ. Çàùèòíàÿ ôóíêöèÿ áåëêîâ ïðîÿâëÿåòñÿ è â ñïîñîáíîñòè ðÿäà áåëêîâ ïëàçìû êðîâè, â ÷àñòíîñòè ôèáðèíîãåíà, ê ñâåðòûâàíèþ. Â ðåçóëüòàòå ñâåðòûâàíèÿ ôèáðèíîãåíà îáðàçóåòñÿ ñãóñòîê êðîâè, ïðåäîõðàíÿþùèé îò ïîòåðè êðîâè ïðè ðàíåíèÿõ. Ñîêðàòèòåëüíàÿ ôóíêöèÿ. Â àêòå ìûøå÷íîãî ñîêðàùåíèÿ è ðàññëàáëåíèÿ ó÷àñòâóåò ìíîæåñòâî áåëêîâûõ âåùåñòâ. Îäíàêî ãëàâíóþ ðîëü â ýòèõ æèçíåííî âàæíûõ ïðîöåññàõ èãðàþò àêòèí è ìèîçèí—ñïåöèôè÷åñêèå áåëêè ìûøå÷íîé òêàíè. Ñîêðàòèòåëüíàÿ ôóíêöèÿ ïðèñóùà íå òîëüêî ìûøå÷íûì áåëêàì, íî è áåëêàì öèòîñêåëåòà, ÷òî îáåñïå÷èâàåò òîí÷àéøèå ïðîöåññû æèçíåäåÿòåëüíîñòè êëåòîê (ðàñõîæäåíèå õðîìîñîì â ïðîöåññå ìèòîçà). Ñòðóêòóðíàÿ ôóíêöèÿ. Áåëêè, âûïîëíÿþùèå ñòðóêòóðíóþ (îïîðíóþ) ôóíêöèþ, çàíèìàþò ïî êîëè÷åñòâó ïåðâîå ìåñòî ñðåäè äðóãèõ áåëêîâ òåëà ÷åëîâåêà. Ñðåäè íèõ âàæíåéøóþ ðîëü èãðàþò ôèáðèëëÿðíûå áåëêè, â ÷àñòíîñòè êîëëàãåí â ñîåäèíèòåëüíîé òêàíè, êåðàòèí â âîëîñàõ, íîãòÿõ, êîæå, ýëàñòèí â ñîñóäèñòîé ñòåíêå è äð. Áîëüøîå çíà÷åíèå èìåþò êîìïëåêñû áåëêîâ ñ óãëåâîäàìè â ôîðìèðîâàíèè ðÿäà ñåêðåòîâ: ìóêîèäîâ, ìóöèíà è ò.ä. Â êîìïëåêñå ñ ëèïèäàìè (â ÷àñòíîñòè, ñ ôîñôîëèïèäàìè) áåëêè ó÷àñòâóþò â îáðàçîâàíèè áèîìåìáðàí êëåòîê. Ãîðìîíàëüíàÿ ôóíêöèÿ. Îáìåí âåùåñòâ â îðãàíèçìå ðåãóëèðóåòñÿ ðàçíîîáðàçíûìè ìåõàíèçìàìè. Â ýòîé ðåãóëÿöèè âàæíîå ìåñòî çàíèìàþò ãîðìîíû, ñèíòåçèðóåìûå íå òîëüêî â æåëåçàõ âíóòðåííåé ñåêðåöèè, íî è âî ìíîãèõ äðóãèõ êëåòêàõ îðãàíèçìà (ñì. äàëåå). Ðÿä ãîðìîíîâ ïðåäñòàâëåí áåëêàìè èëè ïîëèïåïòèäàìè, íàïðèìåð ãîðìîíû ãèïîôèçà, ïîäæåëóäî÷íîé æåëåçû è äð. Íåêîòîðûå ãîðìîíû ÿâëÿþòñÿ ïðîèçâîäíûìè àìèíîêèñëîò. Ïèòàòåëüíàÿ (ðåçåðâíàÿ) ôóíêöèÿ. Ýòó ôóíêöèþ âûïîëíÿþò òàê íàçûâàåìûå ðåçåðâíûå áåëêè, ÿâëÿþùèåñÿ èñòî÷íèêàìè ïèòàíèÿ äëÿ ïëîäà, íàïðèìåð áåëêè ÿéöà (îâàëüáóìèíû). Îñíîâíîé áåëîê ìîëîêà (êàçåèí) òàêæå âûïîëíÿåò ãëàâíûì îáðàçîì ïèòàòåëüíóþ ôóíêöèþ. Ðÿä äðóãèõ áåëêîâ èñïîëüçóåòñÿ â îðãàíèçìå â êà÷åñòâå èñòî÷íèêà àìèíîêèñëîò, êîòîðûå â ñâîþ î÷åðåäü ÿâëÿþòñÿ ïðåäøåñòâåííèêàìè áèîëîãè÷åñêè àêòèâíûõ âåùåñòâ, ðåãóëèðóþùèõ ïðîöåññû ìåòàáîëèçìà. Ìîæíî íàçâàòü åùå íåêîòîðûå äðóãèå æèçíåííî âàæíûå ôóíêöèè áåëêîâ. Ýòî, â ÷àñòíîñòè, ýêñïðåññèÿ ãåíåòè÷åñêîé èíôîðìàöèè, ãåíåðèðîâàíèå è ïåðåäà÷à íåðâíûõ èìïóëüñîâ, ñïîñîáíîñòü ïîääåðæèâàòü îíêîòè÷åñêîå äàâëåíèå â êëåòêàõ è êðîâè, áóôåðíûå ñâîéñòâà, ïîääåðæèâàþùèå ôèçèîëîãè÷åñêîå çíà÷åíèå ðÍ âíóòðåííåé ñðåäû, è äð. Òàêèì îáðàçîì, èç ýòîãî äàëåêî íå ïîëíîãî ïåðå÷íÿ îñíîâíûõ ôóíêöèé áåëêîâ âèäíî, ÷òî óêàçàííûì áèîïîëèìåðàì ïðèíàäëåæèò èñêëþ÷èòåëüíàÿ è ðàçíîñòîðîííÿÿ ðîëü â æèâîì îðãàíèçìå. Åñëè ïîïûòàòüñÿ âûäåëèòü ãëàâíîå, ðåøàþùåå ñâîéñòâî, êîòîðîå îáåñïå÷èâàåò ìíîãîãðàííîñòü áèîëîãè÷åñêèõ ôóíêöèé áåëêîâ, òî ñëåäîâàëî áû íàçâàòü ñïîñîáíîñòü áåëêîâ ñòðîãî èçáèðàòåëüíî, ñïåöèôè÷åñêè ñîåäèíÿòüñÿ ñ øèðîêèì êðóãîì ðàçíîîáðàçíûõ âåùåñòâ. Â ÷àñòíîñòè, ýòà âûñîêàÿ ñïåöèôè÷íîñòü áåëêîâ (ñðîäñòâî) îáåñïå÷èâàåò âçàèìîäåéñòâèå ôåðìåíòîâ ñ ñóáñòðàòàìè, àíòèòåë ñ àíòèãåíàìè, òðàíñïîðòíûõ áåëêîâ êðîâè ñ ïåðåíîñèìûìè ìîëåêóëàìè äðóãèõ âåùåñòâ è ò.ä. Ýòî âçàèìîäåéñòâèå îñíîâàíî íà ïðèíöèïå áèîñïå- öèôè÷åñêîãî óçíàâàíèÿ, çàâåðøàþùåãîñÿ ñâÿçûâàíèåì ôåðìåíòà ñ ñîîòâåòñòâóþùåé ìîëåêóëîé ñóáñòðàòà, ÷òî ñîäåéñòâóåò ïðîòåêàíèþ õèìè÷åñêîé ðåàêöèè. Âûñîêîé ñïåöèôè÷íîñòüþ äåéñòâèÿ íàäåëåíû òàêæå áåëêè, êîòîðûå ó÷àñòâóþò â òàêèõ ïðîöåññàõ, êàê äèôôåðåíöèðîâêà è äåëåíèå êëåòîê, ðàçâèòèå æèâûõ îðãàíèçìîâ, îïðåäåëÿÿ èõ áèîëîãè÷åñêóþ èíäèâèäóàëüíîñòü. ÑÎÄÅÐÆÀÍÈÅ ÁÅËÊÎÂ Â ÎÐÃÀÍÀÕ È ÒÊÀÍßÕ Íàèáîëåå áîãàòû áåëêîâûìè âåùåñòâàìè òêàíè è îðãàíû æèâîòíûõ. Èñòî÷íèêîì áåëêà ÿâëÿþòñÿ òàêæå ìèêðîîðãàíèçìû è ðàñòåíèÿ. Áîëüøèíñòâî áåëêîâ õîðîøî ðàñòâîðèìî â âîäå. Íåêîòîðûå îðãàíè÷åñêèå âåùåñòâà, âûäåëåííûå èç õðÿùà, âîëîñ, íîãòåé, ðîãîâ, êîñòíîé òêàíè è íåðàñòâîðèìûå â âîäå, òàêæå áûëè îòíåñåíû ê áåëêàì, ïîñêîëüêó ïî ñâîåìó õèìè÷åñêîìó ñîñòàâó îêàçàëèñü áëèçêè ê áåëêàì ìûøå÷íîé òêàíè, ñûâîðîòêè êðîâè, ÿéöà. Â ìûøöàõ, ëåãêèõ, ñåëåçåíêå, ïî÷êàõ íà äîëþ áåëêîâ ïðèõîäèòñÿ áîëåå 70—80% îò ñóõîé ìàññû, à âî âñåì òåëå ÷åëîâåêà—45% îò ñóõîé ìàññû (òàáë. 1.1) . Â îòëè÷èå îò æèâîòíûõ òêàíåé â ðàñòåíèÿõ ñîäåðæèòñÿ çíà÷èòåëüíî ìåíüøå áåëêîâ (òàáë. 1.2). Îðãàíû è òêàíè îò ñóõîé ìàññû îò îáùåãî êîëè-÷åñòâà áåëêà òåëà Îðãàíû è òêàíè îò ñóõîé ìàññû îò îáùå-ãî êîëè-÷åñòâà áåëêà òåëà Êîæà 63 11,5 Ïî÷êè 72 0,5 Êîñòè (òâåðäûå òêàíè) 20 18,7 Ïîäæåëóäî÷íàÿ 47 0,1 Çóáû (òâåðäûå òêàíè) 18 0,1 æåëåçà Ïîïåðå÷íîïîëîñàòûå 80 34,7 Ïèùåâàðèòåëüíûé 63 1,8 ìûøöû òðàêò Ìîçã è íåðâíàÿ òêàíü 45 2,0 Æèðîâàÿ òêàíü 14 6,4 Ïå÷åíü 57 3,6 Îñòàëüíûå òêàíè: Ñåðäöå 60 0,7 æèäêèå 85 1,4 Ëåãêèå 82 3,7 ïëîòíûå 54 14,6 Ñåëåçåíêà 84 0,2 Âñå òåëî 45 100 Òàáëèöà 1.1. Ñîäåðæàíèå áåëêîâ â îðãàíàõ è òêàíÿõ ÷åëîâåêà Ñîäåðæàíèå áåëêîâ, % Ñîäåðæàíèå áåëêîâ, % Òàáëèöà 1.2. Ñîäåðæàíèå áåëêîâ â îðãàíàõ æèâîòíûõ è â ðàñòåíèÿõ Îðãàíû æèâîòíûõ Ñîäåðæàíèå áåëêîâ, % îò ìàññû ñâåæåé òêàíè Îðãàíû ðàñòåíèé Ñîäåðæàíèå áåëêîâ, % îò ìàññû ñâåæåé òêàíè Ìûøöû 18-23 Ñåìåíà 10-13 Ïå÷åíü 18-19 Ñòåáëè 1,5-3,0 Ñåëåçåíêà 17-18 Ëèñòüÿ 1,2-3,0 Ïî÷êè 16-18 Êîðíè 0,5-3,0 Ëåãêèå 14-15 Ôðóêòû 0,3-1,0 Ìîçã 7-9 Äëÿ èçó÷åíèÿ õèìè÷åñêîãî ñîñòàâà, ñòðîåíèÿ è ñâîéñòâ áåëêîâ èõ îáû÷íî âûäåëÿþò èëè èç òêàíåé, èëè èç êóëüòèâèðóåìûõ êëåòîê, èëè áèîëîãè÷åñêèõ æèäêîñòåé, íàïðèìåð ñûâîðîòêè êðîâè, ìîëîêà, ìûøö, ïå÷åíè, êîæè è äð. Ýëåìåíòíûé ñîñòàâ áåëêîâ â ïåðåñ÷åòå íà ñóõîå âåùåñòâî ïðåäñòàâëåí 50—54% óãëåðîäà, 21—23% êèñëîðîäà, 6,5—7,3% âîäîðîäà, 15—17% àçîòà è äî 0,5% ñåðû. Â ñîñòàâå íåêîòîðûõ áåëêîâ ïðèñóòñòâóþò â íåáîëüøèõ êîëè÷åñòâàõ ôîñôîð, æåëåçî, ìàðãàíåö, ìàãíèé, éîä è äð. Òàêèì îáðàçîì, ïîìèìî óãëåðîäà, êèñëîðîäà è âîäîðîäà, âõîäÿùèõ â ñîñòàâ ïî÷òè âñåõ îðãàíè÷åñêèõ ïîëèìåðíûõ ìîëåêóë, îáÿçàòåëüíûì êîìïîíåíòîì áåëêîâ ÿâëÿåòñÿ àçîò, â ñâÿçè ñ ÷åì áåëêè ïðèíÿòî îáîçíà÷àòü êàê àçîòñîäåðæàùèå îðãàíè÷åñêèå âåùåñòâà. Ñîäåðæàíèå àçîòà áîëåå èëè ìåíåå ïîñòîÿííî âî âñåõ áåëêàõ (â ñðåäíåì 16%), ïîýòîìó èíîãäà îïðåäåëÿþò êîëè÷åñòâî áåëêà â áèîëîãè÷åñêèõ îáúåêòàõ ïî ñîäåðæàíèþ áåëêîâîãî àçîòà. ÌÅÒÎÄÛ ÂÛÄÅËÅÍÈß È Î×ÈÑÒÊÈ ÁÅËÊÎÂ Äëÿ ïîäðîáíîãî èññëåäîâàíèÿ ôèçèêî-õèìè÷åñêèõ è áèîëîãè÷åñêèõ ñâîéñòâ áåëêîâ, à òàêæå äëÿ èçó÷åíèÿ èõ õèìè÷åñêîãî ñîñòàâà è ñòðóêòóðû íåïðåìåííûì óñëîâèåì ÿâëÿåòñÿ ïîëó÷åíèå áåëêîâ èç ïðèðîäíûõ èñòî÷íèêîâ â õèìè÷åñêè ÷èñòîì, ãîìîãåííîì ñîñòîÿíèè. Ïîñëåäîâàòåëüíîñòü îïåðàöèé ïî âûäåëåíèþ áåëêîâ îáû÷íî ñîñòîèò â ñëåäóþùåì: èçìåëü÷åíèå áèîëîãè÷åñêîãî ìàòåðèàëà (ãîìîãåíèçàöèÿ); èçâëå÷åíèå áåëêîâ, òî÷íåå, ïåðåâîä áåëêîâ â ðàñòâîðåííîå ñîñòîÿíèå (ýêñòðàêöèÿ); âûäåëåíèå èññëåäóåìîãî áåëêà èç ñìåñè äðóãèõ áåëêîâ, ò.å. î÷èñòêà è ïîëó÷åíèå èíäèâèäóàëüíîãî áåëêà. Áåëêîâûå âåùåñòâà âåñüìà ÷óâñòâèòåëüíû ê ïîâûøåíèþ òåìïåðàòóðû è äåéñòâèþ ìíîãèõ õèìè÷åñêèõ ðåàãåíòîâ (îðãàíè÷åñêèå ðàñòâîðèòåëè, êèñëîòû, ùåëî÷è). Ïîýòîìó îáû÷íûå ìåòîäû îðãàíè÷åñêîé õèìèè, ïðèìåíÿåìûå äëÿ âûäåëåíèÿ òîãî èëè èíîãî âåùåñòâà èç ñìåñè (íàãðåâàíèå, ïåðåãîíêà, âîçãîíêà, êðèñòàëëèçàöèÿ è äð.), â äàííîì ñëó÷àå íåïðèåìëåìû. Áåëêè â ýòèõ óñëîâèÿõ ïîäâåðãàþòñÿ äåíàòóðàöèè, ò.å. òåðÿþò íåêîòîðûå ñóùåñòâåííûå ïðèðîäíûå (íàòèâíûå) ñâîéñòâà, â ÷àñòíîñòè ðàñòâîðèìîñòü, áèîëîãè÷åñêóþ àêòèâíîñòü. Ðàçðàáîòàíû ýôôåêòèâíûå ìåòîäû âûäåëåíèÿ áåëêîâ â «ìÿãêèõ» óñëîâèÿõ, ïðè íèçêîé òåìïåðàòóðå (íå âûøå 4°Ñ), ñ ïðèìåíåíèåì ùàäÿùèõ íàòèâíóþ ñòðóêòóðó õèìè÷åñêèõ ðåàãåíòîâ. Ãîìîãåíèçàöèÿ áèîëîãè÷åñêîãî ìàòåðèàëà Ïåðåä âûäåëåíèåì áåëêîâ èç áèîëîãè÷åñêèõ îáúåêòîâ (îðãàíû è òêàíè æèâîòíûõ, ìèêðîîðãàíèçìû, ðàñòåíèÿ) èññëåäóåìûé ìàòåðèàë òùàòåëüíî èçìåëü÷àþò äî ãîìîãåííîãî ñîñòîÿíèÿ, ò.å. ïîäâåðãàþò äåçèíòåãðàöèè âïëîòü äî ðàçðóøåíèÿ êëåòî÷íîé ñòðóêòóðû. Ýòó ïðîöåäóðó, íàçûâàåìóþ ãîìîãåíèçàöèåé, ïðîâîäÿò ïðè ïîìîùè íîæåâûõ ãîìîãåíèçàòîðîâ òèïà Óîððèíãà èëè ïåñòèêîâîãî ãîìîãåíèçàòîðà Ïîòòåðà—Ýëüâåãåéìà. Äëÿ âûäåëåíèÿ ðÿäà áåëêîâ èç ïëîòíûõ æèâîòíûõ è ðàñòèòåëüíûõ îáúåêòîâ ÷àñòî èñïîëüçóþò âàëêîâûå èëè øàðîâûå ìåëüíèöû (ðèñ. 1.1). Óñïåøíî ïðèìåíÿåòñÿ òàêæå ìåòîä ïîïåðåìåííîãî çàìîðàæèâàíèÿ è îòòàèâàíèÿ òêàíè, â îñíîâå äåéñòâèÿ êîòîðîãî ëåæèò ðàçðóøåíèå êëåòî÷íîé îáîëî÷êè, âûçâàííîå êðèñòàëëàìè ëüäà. Äëÿ äåçèíòåãðàöèè òêàíåé èñïîëüçóþò òàêæå óëüòðàçâóê, ïðåññ-ìåòîäû (çàìîðîæåííûé áèîìàòåðèàë ïðîïóñêàþò ÷åðåç ìåëü÷àéøèå îòâåðñòèÿ ñòàëüíîãî ïðåññà ïîä âûñîêèì äàâëåíèåì) è ìåòîä «àçîòíîé áîìáû», ïðè êîòîðîì êëåòêè (â ÷àñòíîñòè, ìèêðîáíûå) ñíà÷àëà íàñûùàþò àçîòîì ïîä âûñîêèì äàâëåíèåì, çàòåì ðåçêî ñáðàñûâàþò äàâëå- íèå—âûäåëÿþùèéñÿ ãàçîîáðàçíûé àçîò êàê áû «âçðûâàåò» êëåòêè. Ýêñòðàêöèÿ áåëêîâ Ñîâðåìåííûå ìåòîäû èçìåëü÷åíèÿ òêàíåé îáû÷íî ñî÷åòàþò ñ îäíîâðåìåííîé ýêñòðàêöèåé áåëêîâ èç ãîìîãåíàòîâ òêàíåé. Áîëüøèíñòâî áåëêîâ òêàíåé õîðîøî ðàñòâîðèìî â 8—10% ðàñòâîðàõ ñîëåé. Ïðè ýêñòðàêöèè áåëêîâ øèðîêî ïðèìåíÿþò ðàçëè÷íûå áóôåðíûå ñìåñè ñ îïðåäåëåííûìè çíà÷åíèÿìè ðÍ ñðåäû, îðãàíè÷åñêèå ðàñòâîðèòåëè, à òàêæå íåèîííûå äåòåðãåíòû — âåùåñòâà, ðàçðóøàþùèå ãèäðîôîáíûå âçàèìîäåéñòâèÿ ìåæäó áåëêàìè è ëèïèäàìè è ìåæäó áåëêîâûìè ìîëåêóëàìè. Èç îðãàíè÷åñêèõ ñîåäèíåíèé, ïîìèìî äàâíî ïðèìåíÿåìûõ âîäíûõ ðàñòâîðîâ ãëèöåðèíà, øèðîêî èñïîëüçóþò (îñîáåííî äëÿ ñîëþáèëèçàöèè) ñëàáûå ðàñòâîðû ñàõàðîçû. Íà ðàñòâîðèìîñòü áåëêîâ ïðè ýêñòðàêöèè áîëüøîå âëèÿíèå îêàçûâàåò ðÍ ñðåäû, ïîýòîìó â áåëêîâîé õèìèè ïðèìåíÿþò ôîñôàòíûå, öèòðàòíûå, áîðàòíûå áóôåðíûå ñìåñè ñî çíà÷åíèÿìè ðÍ îò êèñëûõ äî ñëàáîùåëî÷íûõ, êîòîðûå ñïîñîáñòâóþò êàê ðàñòâîðåíèþ, òàê è ñòàáèëèçàöèè áåëêîâ. Îñîáåííî øèðîêîå ðàñïðîñòðàíåíèå ïîëó÷èëè òðèñ-áóôåðíûå ñèñòåìû, ïðåäñòàâëÿþùèå ñîáîé ñìåñè 0,2 Ì ðàñòâîðà òðèñ-(îêñèìåòèë)-àìèíîìåòàíà (ÍÎÑÍ2)3ÑÍÍ2 (ñîêðàùåííî îáîçíà÷àþò «òðèñ») ñ 0,1 Ì ðàñòâîðîì õëîðîâîäîðîäíîé êèñëîòû â ðàçíûõ ñîîòíîøåíèÿõ. Äëÿ âûäåëåíèÿ áåëêîâ ñûâîðîòêè êðîâè èñïîëüçóþò ñïîñîáû èõ îñàæäåíèÿ ýòàíîëîì (ñì. ìåòîä Êîíà), àöåòîíîì, áóòàíîëîì è èõ êîìáèíàöèè. Ïî÷òè âñå îðãàíè÷åñêèå ðàñòâîðèòåëè ðàçðûâàþò áåëîê-ëèïèäíûå ñâÿçè, ñïîñîáñòâóÿ ëó÷øåé ýêñòðàêöèè áåëêîâ. Äëÿ ïîëó÷åíèÿ èç áèîëîãè÷åñêîãî ìàòåðèàëà áåëêîâ â ÷èñòîì, ãîìîãåííîì, ñîñòîÿíèè ïðèìåíÿþò ðàçëè÷íûå äåòåðãåíòû, ñïîñîáñòâóþùèå ðàñùåïëåíèþ áåëîê-ëèïèäíûõ êîìïëåêñîâ è ðàçðûâó áåëîê-áåëêîâûõ ñâÿçåé . Â ÷àñòíîñòè, äëÿ îñâîáîæäåíèÿ áåëêîâ (ôåðìåíòîâ), ïðî÷íî ñâÿçàííûõ ñ áèîìåìáðàíàìè ìèòîõîíäðèé èëè äðóãèõ ñóáêëåòî÷íûõ ñòðóêòóð, ïðèìåíÿþò òðèòîí Õ-100, äîäåöèëñóëüôàò íàòðèÿ è äåçîêñèõîëàò íàòðèÿ. ÑÍ3—(ÑÍ2)10—ÑÍ2—Î—5Î3Ûà Äîäåöèëñóëüôàò íàòðèÿ Ñ8Í17—(Ñ6Í4)—Î—(ÑÍ2—ÑÍÎÍ)þÍ Òðèòîí Õ-100 Ðèñ. 1.1. Ëàáîðàòîðíîå îáîðóäîâàíèå. á à—ïåñòèêîâûé ðó÷íîé ãîìîãåíèçàòîð: 1 — ïåñòèê, 2—êîðïóñ, 3 — ìîòîð, 4—øòàòèâ; ìåõàíè÷åñêèé ãîìîãåíèçàòîð (á) è øàðîâàÿ ìåëüíèöà (â): 1 — êîðïóñ ñ ýëåêòðîäâèãàòåëåì è ïóñêîâûì óñòðîéñòâîì, 2 — êàìåðà äëÿ èçìåëü÷åíèÿ ìàòåðèàëà. Ñëåäóåò, îäíàêî, èìåòü â âèäó, ÷òî äåòåðãåíòû, âûçûâàÿ ðàçðûâ áåëîê- áåëêîâûõ ñâÿçåé, ðàçðóøàþò îëèãîìåðíóþ (÷åòâåðòè÷íóþ) ñòðóêòóðó áåëêîâ. Ôðàêöèîíèðîâàíèå è î÷èñòêà áåëêîâ Ïîñëå äîñòèæåíèÿ ïîëíîé ýêñòðàêöèè áåëêîâ, ò.å. ïåðåâîäà áåëêîâ â ðàñòâîðåííîå ñîñòîÿíèå, ïðèñòóïàþò ê ðàçäåëåíèþ — ôðàêöèîíèðîâàíèþ ñìåñè áåëêîâ íà èíäèâèäóàëüíûå áåëêè. Äëÿ ýòîãî ïðèìåíÿþò ðàçíîîáðàçíûå ìåòîäû: âûñàëèâàíèå, òåïëîâóþ äåíàòóðàöèþ, îñàæäåíèå îðãàíè÷åñêèìè ðàñòâîðèòåëÿìè, õðîìàòîãðàôèþ, ýëåêòðîôîðåç, ðàñïðåäåëåíèå â äâóõôàçíûõ ñèñòåìàõ, êðèñòàëëèçàöèþ è äð. Ðàñòâîðåíèå áåëêîâ â âîäå ñâÿçàíî ñ ãèäðàòàöèåé êàæäîé ìîëåêóëû, ÷òî ïðèâîäèò ê îáðàçîâàíèþ âîêðóã áåëêîâîé ãëîáóëû âîäíûõ (ãèäðàòíûõ) îáîëî÷åê, ñîñòîÿùèõ èç îðèåíòèðîâàííûõ â îïðåäåëåííîé ôîðìå â ïðîñòðàíñòâå ìîëåêóë âîäû. Ïî õèìè÷åñêèì è ôèçè÷åñêèì ñâîéñòâàì âîäà, âõîäÿùàÿ â ñîñòàâ ãèäðàòíîé îáîëî÷êè, îòëè÷àåòñÿ îò ÷èñòîãî ðàñòâîðèòåëÿ. Â ÷àñòíîñòè, òåìïåðàòóðà çàìåðçàíèÿ åå ñîñòàâëÿåò —40°Ñ. Â ýòîé âîäå õóæå ðàñòâîðÿþòñÿ ñàõàðà, ñîëè è äðóãèå âåùåñòâà. Ðàñòâîðû áåëêîâ îòëè÷àþòñÿ êðàéíåé íåóñòîé÷èâîñòüþ, è ïîä äåéñòâèåì ðàçíîîáðàçíûõ ôàêòîðîâ, íàðóøàþùèõ ãèäðàòàöèþ, áåëêè ëåãêî âûïàäàþò â îñàäîê. Ïîýòîìó ïðè äîáàâëåíèè ê ðàñòâîðó áåëêà ëþáûõ âîäîîòíèìàþùèõ ñðåäñòâ (ñïèðò, àöåòîí, êîíöåíòðèðîâàííûå ðàñòâîðû íåéòðàëüíûõ ñîëåé ùåëî÷íûõ ìåòàëëîâ), à òàêæå ïîä âëèÿíèåì ôèçè÷åñêèõ ôàêòîðîâ (íàãðåâàíèå, îáëó÷åíèå è äð.) íàáëþäàþòñÿ äåãèäðàòàöèÿ ìîëåêóë áåëêà è èõ âûïàäåíèå â îñàäîê. Âûñàëèâàíèå. Ïðè äîáàâëåíèè ðàñòâîðîâ ñîëåé ùåëî÷íûõ è ùåëî÷íîçåìåëüíûõ ìåòàëëîâ ïðîèñõîäèò îñàæäåíèå áåëêîâ èç ðàñòâîðà. Îáû÷íî áåëîê íå òåðÿåò ñïîñîáíîñòè ðàñòâîðÿòüñÿ âíîâü â âîäå ïîñëå óäàëåíèÿ ñîëåé ìåòîäàìè äèàëèçà èëè ãåëüõðîìàòîãðàôèè. Âûñàëèâàíèåì áåëêîâ îáû÷íî ïîëüçóþòñÿ â êëèíè÷åñêîé ïðàêòèêå ïðè àíàëèçå áåëêîâ ñûâîðîòêè êðîâè è äðóãèõ áèîëîãè÷åñêèõ æèäêîñòåé, à òàêæå â ïðåïàðàòèâíîé ýíçèìî- ëîãèè äëÿ ïðåäâàðèòåëüíîãî îñàæäåíèÿ è óäàëåíèÿ áàëëàñòíûõ áåëêîâ èëè âûäåëåíèÿ èññëåäóåìîãî ôåðìåíòà. Ðàçëè÷íûå áåëêè âûñàëèâàþòñÿ èç ðàñòâîðîâ ïðè ðàçíûõ êîíöåíòðàöèÿõ íåéòðàëüíûõ ðàñòâîðîâ ñóëüôàòà àììîíèÿ. Ïîýòîìó ìåòîä íàøåë øèðîêîå ïðèìåíåíèå â êëèíèêå äëÿ ðàçäåëåíèÿ ãëîáóëèíîâ (âûïàäàþò â îñàäîê ïðè 50% íàñûùåíèè) è àëüáóìèíîâ (âûïàäàþò ïðè 100% íàñûùåíèè). Íà âåëè÷èíó âûñàëèâàíèÿ áåëêîâ îêàçûâàþò âëèÿíèå íå òîëüêî ïðèðîäà è êîíöåíòðàöèÿ ñîëè, íî è ðÍ ñðåäû è òåìïåðàòóðà. Ñ÷èòàþò, ÷òî ãëàâíóþ ðîëü ïðè ýòîì èãðàåò âàëåíòíîñòü èîíîâ. Äåéñòâèå ðàçíûõ èîíîâ ïðèíÿòî ñðàâíèâàòü íå ïî ìîëÿðíîé êîíöåíòðàöèè ñîëè, à ïî òàê íàçûâàåìîé èîííîé ñèëå ô), êîòîðàÿ ðàâíà ïîëîâèíå ñóììû ïðîèçâåäåíèé êîíöåíòðàöèè êàæäîãî èîíà (ñ) íà êâàäðàò åãî âàëåíòíîñòè (V): Áîëåå òîíêîå ðàçäåëåíèå áåëêîâ ïëàçìû êðîâè ÷åëîâåêà íà ôðàêöèè äîñòèãàåòñÿ ïðè èñïîëüçîâàíèè ðàçëè÷íûõ êîíöåíòðàöèé ýòàíîëà ïðè íèçêîé òåìïåðàòóðå (îò —3 äî —5°Ñ) ïî ìåòîäó Êîíà (ðèñ. 1.2). Â ýòèõ óñëîâèÿõ áåëêè ñîõðàíÿþò ñâîè íàòèâíûå ñâîéñòâà. Óêàçàííûì ìåòîäîì ÷àñòî ïîëüçóþòñÿ äëÿ ïîëó÷åíèÿ îòäåëüíûõ ôðàêöèé êðîâè, èñïîëüçóåìûõ â êà÷åñòâå êðîâåçàìåíèòåëåé. Ðèñ. 1.2. Äèàãðàììà ôðàêöèîíèðîâàíèÿ áåëêîâ ïëàçìû êðîâè ÷åëîâåêà ýòàíîëîì (ïî ìåòîäó Êîíà). Â ïîñëåäíåå âðåìÿ íàèáîëüøåå ðàñïðîñòðàíåíèå ïîëó÷èëè õðîìàòîãðàôè÷åñêèå è ýëåêòðîôîðåòè÷åñêèå ìåòîäû ðàçäåëåíèÿ áåëêîâ. Õðîìàòîãðàôèÿ. Ïðèíöèï õðîìàòîãðàôèè, ðàçðàáîòàííûé â 1903 ã. ðóññêèì ó÷åíûì Ì. Ñ. Öâåòîì, îñíîâàí íà ñïîñîáíîñòè ïèãìåíòîâ (èëè ëþáûõ äðóãèõ îêðàøåííûõ è íåîêðàøåííûõ âåùåñòâ) ñïåöèôè÷åñêè àäñîðáèðîâàòüñÿ íà àäñîðáåíòå, çàêëþ÷åííîì â êîëîíêå . Â ðåçóëüòàòå ïðîèñõîäèò ðàçäåëåíèå àíàëèçèðóåìûõ âåùåñòâ è èõ êîíöåíòðèðîâàíèå â ñòðîãî îïðåäåëåííîì ñëîå àäñîðáåíòà. Çàòåì ÷åðåç êîëîíêó ïðîïóñêàþò ïîäõîäÿùèå ýëþåíòû, êîòîðûå îñëàáëÿþò ñèëû àäñîðáöèè è âûíîñÿò ñ òîêîì ðàñòâîðà èíäèâèäóàëüíûå âåùåñòâà. Ïîñëåäíèå ïîñëåäîâàòåëüíî ñîáèðàþò â êîëëåêòîðå ôðàêöèé (ïðèíöèï ñîðáöèè-äå-ñîðáöèè). ×ðåçâû÷àéíî ýôôåêòèâíûì ñðåäñòâîì ôðàêöèîíèðîâàíèÿ áåëêîâ èç ñìåñè îêàçàëàñü êîëîíî÷íàÿ õðîìàòîãðàôèÿ ñ ãèäðîêñèëàïàòèòîì, ðàçëè÷íûìè èîíîîáìåííûìè ñìîëàìè è ïðîèçâîäíûìè öåëëëþëîçû â êà÷åñòâå íîñèòåëåé. Ïðè âûäåëåíèè è î÷èñòêå áåëêîâ èñïîëüçóþò ÷åòûðå îñíîâíûõ òèïà õðîìàòîãðàôèè: àäñîðáöèîííóþ, ðàñïðåäåëèòåëüíóþ, èîíîîáìåííóþ è àôôèííóþ (õðîìàòîãðàôèÿ ïî ñðîäñòâó) — â ñîîòâåòñòâèè ñ ðàçíûìè ôèçè÷åñêèìè è õèìè÷åñêèìè ìåõàíèçìàìè, ëåæàùèìè â îñíîâå êàæäîãî èç íèõ. Õðîìàòîãðàôèÿ øèðîêî ïðèìåíÿåòñÿ íå òîëüêî äëÿ âûäåëåíèÿ áåëêîâ, íî è äëÿ ðàçäåëåíèÿ ìíîæåñòâà äðóãèõ îðãàíè÷åñêèõ è íåîðãàíè÷åñêèõ âåùåñòâ, âõîäÿùèõ â ñîñòàâ æèâûõ îðãàíèçìîâ. Àäñîðáöèîííàÿ õðîìàòîãðàôèÿ. Ðàçäåëåíèå êîìïîíåíòîâ ñìåñè (îáðàçöà) îñíîâàíî íà èõ ðàçëè÷íîé ñîðáèðóåìîñòè íà òâåðäîì àäñîðáåíòå. Â êà÷åñòâå àäñîðáåíòîâ èñïîëüçóþò àêòèâèðîâàííûé äðåâåñíûé óãîëü, ãåëü ôîñôàòà êàëüöèÿ, îêñèäû àëþìèíèÿ èëè êðåìíèÿ. Àäñîðáåíò â âèäå ñóñïåíçèè ñ ðàñòâîðèòåëåì (÷àùå âñåãî áóôåðíûì ðàñòâîðîì) âíîñÿò â ñòåêëÿííóþ âåðòèêàëüíóþ òðóáêó (êîëîíêó) è ðàâíîìåðíî â íåé óïàêîâûâàþò. Îáðàçåö â íåáîëüøîì îáúåìå ðàñòâîðèòåëÿ íàíîñÿò íà êîëîíêó — Ýëþåíò^ êîìïîíåíòû ðàçäåëÿåìîé ñìåñè àäñîðáèðóþòñÿ íà àäñîðáåíòå. Çàòåì ïðèñòóïàþò ê ñòàäèè îñâîáîæäåíèÿ—äåñîðáöèè êîìïîíåíòîâ èç êîëîíêè, ïðèìåíÿÿ ïîäõîäÿùèå ýëþåíòû (ðèñ. 1.3). Ñáîð ôðàêöèé îñóùåñòâëÿþò ïðè ïîìîùè àâòîìàòè÷åñêîãî êîëëåêòîðà ôðàêöèé. Ðàñïðåäåëèòåëüíàÿ õðîìàòîãðàôèÿ. Â îòëè÷èå îò àäñîðáöèîííîé òâåðäàÿ ôàçà ñëóæèò òîëüêî îïîðîé (îñíîâîé) äëÿ ñòàöèîíàðíîé æèäêîé ôàçû. Îäèí èç òèïîâ ðàñïðåäåëèòåëüíîé õðîìàòîãðàôèè, êàê è àäñîðáöèîííàÿ, îñóùåñòâëÿåòñÿ íà êîëîíêàõ, â êîòîðûõ â êà÷åñòâå ñòàöèîíàðíîé ôàçû ïðèìåíÿþò âëàæíûé êðàõìàë èëè ñèëèêàãåëü. Îáðàçåö ðàñòâîðÿþò â ïîä-õîäÿùåì ðàñòâîðèòåëå, çàòåì íàíîñÿò íà êîëîíêó; ðàçäåëÿåìûå âåùåñòâà, ïîäâåðãàþùèåñÿ ìíîãîêðàòíîìó ðàñïðåäåëåíèþ ìåæäó íåïîäâèæíîé ñòàöèîíàðíîé ôàçîé (âîäíûé ñëîé) è äâèæóùåéñÿ ôàçîé îðãàíè÷åñêîãî ðàñòâîðèòåëÿ, ñ ðàçíîé ñêîðîñòüþ ïåðåìåùàþòñÿ êî äíó êîëîíêè. Ñîáðàííûå ïðè ïîìîùè êîëëåêòîðà ôðàêöèè ïðîáû, ñîäåðæàùèå îäíî âåùåñòâî, ñîåäèíÿþò äëÿ âûäåëåíèÿ ýòîãî âåùåñòâà â ÷èñòîì âèäå. Ðàçíîâèäíîñòüþ ðàñïðåäåëèòåëüíîé õðîìàòîãðàôèè ÿâëÿåòñÿ õðîìàòîãðàôèÿ íà áóìàãå, øèðîêî èñïîëüçóåìàÿ â áèîõèìè÷åñêèõ ëàáîðàòîðèÿõ, â òîì ÷èñëå êëèíè÷åñêèõ, äëÿ ðàçäåëåíèÿ ïåïòèäîâ, àìèíîêèñëîò è äðóãèõ âåùåñòâ (ðèñ. 1.4). Â êà÷åñòâå ñòàöèîíàðíîé ôàçû ïðè ýòîì ñëóæèò âîäà, àäñîðáèðîâàííàÿ öåëëþëîçíûìè öåïÿìè ôèëüòðîâàëüíîé áóìàãè. Îáðàçåö ïîìåùàþò íà îäíîì êîíöå áóìàæíîé ïîëîñû, ýòèì æå êîíöîì áóìàãó ïîãðóæàþò â ïîäõîäÿùóþ ñìåñü îðãàíè÷åñêèõ ðàñòâîðèòåëåé (íàïðèìåð, áóòàíîë—óêñóñíàÿ êèñëîòà—âîäà â îïðåäåëåííûõ ñîîòíîøåíèÿõ). Ïðè äâèæåíèè ðàñòâîðèòåëÿ ïî áóìàãå áëàãîäàðÿ ñèëå êàïèëëÿðíîñòè ïðîèñõîäèò ðàçäåëåíèå êîìïîíåíòîâ ñìåñè. Ïðîÿâëåííóþ õðîìàòîãðàììó âûñóøèâàþò, à ìåñòîïîëîæåíèå êàæäîãî èç ðàçäåëÿåìûõ âåùåñòâ îïðåäåëÿþò õèìè÷åñêèìè èëè ôèçèêî-õèìè÷åñêèìè ìåòîäàìè. Èîíîîáìåííàÿ õðîìàòîãðàôèÿ. Èîíîîáìåííûå ñìîëû ÿâëÿþòñÿ ïîëè-ìåðíûìè îðãàíè÷åñêèìè ñîåäèíåíèÿìè, ñîäåðæàùèìè ôóíêöèîíàëüíûå ãðóïïû, ñïîñîáíûå âîâëåêàòüñÿ â èîííûé îáìåí. Ðàçëè÷àþò ïîëîæèòåëüíî çàðÿæåííûå àíèîíîîáìåííèêè, ïðåäñòàâëåííûå îðãàíè÷åñêèìè îñíîâàíèÿìè è àìèíàìè, è îòðèöàòåëüíî çàðÿæåííûå êàòèîíîîáìåííèêè, ñîäåðæàùèå ôåíîëüíûå, ñóëüôî- èëè êàðáîêñèëüíûå ãðóïïû. Èç ñèëüíî- è ñëàáîîñíîâíûõ àíèîíîîáìåííèêîâ ÷àùå èñïîëüçóþò ïðîèçâîäíûå ïîëèñòèðîëà è öåëëþëîçû, íåñóùèå ôóíêöèîíàëüíûå ãðóïïû: ÑÍà-0-Ñ2Í41×Í(Ñ2Í5)2 Òðèìåòèëàìèíîïîëèñòèðîë Äèýòèëàìèíîýòèëöåëëþëîçà (ÄÝÀÝ-öåëëþëîçà) Àíàëîãè÷íûå ôóíêöèîíàëüíûå ãðóïïû ñîäåðæàò òðèýòèëàìèíîýòèë (ÒÝÀÝ)- è àìèíîýòèë (ÀÝ)-öåëëþëîçû. Êàòèîíîîáìåííèêè ïðåäñòàâëåíû ñóëüôîíèðîâàííûìè ïîëèñòèðîëàìè (ïðîèçâîäíûå âèíèëáåíçîëà èëè äèâèíèëáåíçîëà) è êàðáîêñèìåòèëöåëëþ- ëîçîé, èìåþùèìè ñëåäóþùèå ôóíêöèîíàëüíûå ãðóïïû: ÑÍ2—Î ^ÑÍ^-ÑÎÎ Â çàâèñèìîñòè îò çàðÿäà ðàçäåëÿåìûõ áåëêîâ èñïîëüçóþò ïîäõîäÿùóþ èîíîîáìåííóþ ñìîëó, ñ ôóíêöèîíàëüíûìè ãðóïïàìè êîòîðîé îáìåíèâàåòñÿ è çàäåðæèâàåòñÿ íà êîëîíêå ÷àñòü áåëêîâ, â òî âðåìÿ êàê äðóãèå áåëêè áåñïðåïÿòñòâåííî ýëþèðóþòñÿ ñ êîëîíêè. «Îñàæäåííûå» íà êîëîíêå áåëêè ñíèìàþò ñ êîëîíêè, ïðèìåíÿÿ áîëåå êîíöåíòðèðîâàííûå ñîëåâûå ðàñòâîðû èëè èçìåíÿÿ ðÍ ýëþåíòà. Íîâåéøèå ìåòîäû èîíîîáìåííîé õðîìàòîãðàôèè, â ÷àñòíîñòè âûñîêîýôôåêòèâíàÿ æèäêîñòíàÿ õðîìàòîãðàôèÿ (ÂÝÆÕ), øèðîêî èñïîëüçóþòñÿ â ôàðìàêîëîãèè (ïðè ñîçäàíèè è îïðåäåëåíèè ëåêàðñòâåííûõ âåùåñòâ), â êëèíè÷åñêîé áèîõèìèè (ïðè îïðåäåëåíèè áèîëîãè÷åñêè àêòèâíûõ âåùåñòâ â ôèçèîëîãè÷åñêèõ æèäêîñòÿõ), â áèîòåõíîëîãè÷åñêèõ ïðîöåññàõ è ïðîèçâîäñòâàõ è äðóãèõ îáëàñòÿõ: îíè ïîçâîëÿþò îïðåäåëÿòü âåùåñòâà â íàíî-, ïèêî- è ôåìòàãðàììíûõ êîëè÷åñòâàõ. Àôôèííàÿ õðîìàòîãðàôèÿ (õðîìàòîãðàôèÿ ïî ñðîäñòâó). Îñíîâàíà àôôèííàÿ õðîìàòîãðàôèÿ íà ïðèíöèïå èçáèðàòåëüíîãî âçàèìîäåéñòâèÿ áåëêîâ (èëè äðóãèõ ìàêðîìîëåêóë) ñ çàêðåïëåííûìè (èììîáèëèçîâàííûìè) íà íîñèòåëå ñïåöèôè÷åñêèìè âåùåñòâàìè—ëèãàíäàìè, êîòîðûìè ìîãóò áûòü ñóáñòðàòû èëè êîôåðìåíòû (êîãäà âûäåëÿþò êàêîé-ëèáî ôåðìåíò), àíòèãåíû (èëè àíòèòåëà), ãîðìîíû èëè ðåöåïòîðû è ò. ä. Áëàãîäàðÿ âûñîêîé ñïåöèôè÷íîñòè áåëêîâ ê èììîáèëèçîâàííîìó ëèãàíäó, ñâÿçàííîìó ñ íîñè-òåëåì (êîòîðûì çàïîëíÿþò õðîìàòîãðàôè÷åñêóþ êîëîíêó), ïðèñîåäèíÿåòñÿ òîëüêî îäèí êàêîé-ëèáî áåëîê èç ñìåñè. Ñíÿòèå ñ êîëîíêè ýòîãî áåëêà îñóùåñòâëÿþò ýëþèðîâàíèåì áóôåðíûìè ñìåñÿìè ñ èçìåíåííûì ðÍ èëè èçìåíåííîé èîííîé ñèëîé, à òàêæå ââåäåíèåì â ñîñòàâ ýëþåíòà äåòåðãåíòîâ, îñëàáëÿþùèõ ñâÿçè ìåæäó áåëêàìè è ëèãàíäàìè. Íåñîìíåííûì äîñòîèíñòâîì ìåòîäà ÿâëÿåòñÿ âîçìîæíîñòü îäíîýòàïíî âûäåëèòü çàäàííûé áåëîê èëè äðóãîé áèîïîëèìåð âûñîêîé ñòåïåíè ÷èñòîòû. Ïðè ïîìîùè àôôèííîé õðîìàòîãðàôèè, íàïðèìåð, óäàëîñü ñðàâíèòåëüíî ëåãêî âûäåëèòü î÷èùåííûå ïðåïàðàòû àìèíîàöèë-òÐÍÊ-ñèíòåòàç íà ïîëèàêðèëãèäðàçèäàãàðîâîì ãåëå, ê êîòîðîìó â êà÷åñòâå ëèãàíäîâ áûëè ïðèñîåäèíåíû îïðåäåëåííûå òÐÍÊ (òðàíñïîðòíûå ÐÍÊ). Ãåëü-õðîìàòîãðàôèÿ. Â ïðåïàðàòèâíûõ öåëÿõ, îñîáåííî ïðè î÷èñòêå áåëêîâ îò ïðèìåñåé, øèðîêî èñïîëüçóþò ìåòîä ìîëåêóëÿðíûõ ñèò, èëè ãåëü-õðîìàòîãðàôèþ. Ïðè îáðàáîòêå ýïèõëîðãèäðèíîì ïîëèñàõàðèäà äåê- ñòðàíà îáðàçóþòñÿ ðàçëè÷íîé ñòåïåíè âûðàæåííîñòè ïîïåðå÷íûå ñâÿçè, ïðèâîäÿùèå ê ôîðìèðîâàíèþ êðóïíûõ ãèäðîôèëüíûõ çåðåí, íåðàñòâîðèìûõ â âîäå è íàçûâàåìûõ ñåôàäåêñàìè. Áëàãîäàðÿ áîëüøîìó ñðîäñòâó ê âîäå çåðíà ñèëüíî íàáóõàþò â âîäíîé ñðåäå ñ îáðàçîâàíèåì ãåëÿ, êîòîðûì çàïîëíÿþò õðîìàòîãðàôè÷åñêóþ êîëîíêó. Ðàçäåëåíèå âåùåñòâ ýòèì ìåòîäîì îñíîâàíî íà òîì, ÷òî áîëüøèå ìîëåêóëû íå ïðîíèêàþò âî âíóòðåííþþ âîäíóþ ôàçó ãåëÿ, ÿâëÿþùóþñÿ ñòàöèîíàðíîé, è îñòàþòñÿ ñíàðóæè, äâèãàÿñü âìåñòå ñ ïîäâèæíîé ôàçîé âíèç âäîëü êîëîíêè; íåáîëüøèå ìîëåêóëû, íàïðîòèâ, ñâîáîäíî äèôôóíäèðóþò âíóòðü çåðåí, îáðàçóÿ ðàâíîâåñíóþ ñèñòåìó ìåæäó ïîäâèæíîé è ñòàöèîíàðíîé ôàçàìè, è ñîîòâåòñòâåííî ñ ìåíüøåé ñêîðîñòüþ äâèãàþòñÿ âäîëü êîëîíêè (ðèñ. 1.5). Îáû÷íî ìîìåíò ïîÿâëåíèÿ âåùåñòâ â âûòåêàþùåì èç êîëîíêè ñ ñåôàäåêñîì ýëþåíòå âûðàæàþò ôîðìóëîé: = Ó0 + Ê* V ãäå V — îáúåì ýëþèðóþùåé æèäêîñòè âåùåñòâà ñ äàííûì Ê, ìë; V0 — ñâîáîäíûé îáúåì êîëîíêè èëè îáùèé îáúåì âíåøíåãî ðàñòâîðèòåëÿ (âíå çåðåí ãåëÿ), ìë; V — îáúåì ðàñòâîðèòåëÿ âíóòðè ãåëÿ, ìë; Ê — êîýôôèöèåíò ðàñïðåäåëåíèÿ äëÿ ðàñòâîðåííîãî âåùåñòâà ìåæäó ðàñòâîðèòåëåì âíóòðè çåðåí ãåëÿ è îêðóæàþùèì ðàñòâîðèòåëåì. Åñëè àíàëèçèðóåìóþ ïðîáó, ñîäåðæàùóþ îäíî ðàñòâîðåííîå âåùåñòâî ñ Ê =1 è âòîðîå ñ Ê = 0, âíåñòè â êîëîíêó ñ ñåôàäåêñîì, òî âòîðîå âåùåñòâî ïîÿâèòñÿ â ýëþèðóþùåé æèäêîñòè ñðàçó ïîñëå âûõîäà èç êîëîíêè Ó0, à ïåðâîå—òîëüêî ïîñëå âûõîäà îáúåìà Ó0 + Óã Ïîñêîëüêó ìîëåêóëû áåëêîâ, îáëàäàþùèå áîëüøèìè ìîëåêóëÿðíîé ìàññîé è ðàçìåðàìè, íå äèôôóíäèðóþò âíóòðü çåðåí ñåôàäåêñà, îíè ïåðâûìè âûìûâàþòñÿ èç êîëîíêè ïîñëå âûõîäà ñâîáîäíîãî îáúåìà êîëîíêè Ó0, â òî âðåìÿ êàê âñå îñòàëüíûå âåùåñòâà (âêëþ÷àÿ íèçêîìîëåêóëÿðíûå ïðèìåñè) âûìûâàþòñÿ ïîñëå âûõîäà îáúåìà, ðàâíîãî Ó0 + Ê • Óã Ìåòîä íàøåë øèðîêîå ïðèìåíåíèå â ïðåïàðàòèâíîé ýíçèìîëîãèè. Ñ ïîìîùüþ ñåôàäåêñà ìîæíî ðàçäåëèòü áåëêè ñ ðàçíîé ìîëåêóëÿðíîé ìàññîé. Ýëåêòðîôîðåç. Ìåòîä ñâîáîäíîãî ýëåêòðîôîðåçà, äåòàëüíî ðàçðàáîòàííûé ëàóðåàòîì Íîáåëåâñêîé ïðåìèè À. Òèçåëèóñîì, îñíîâàí íà ðàçëè÷èè â ñêîðîñòè äâèæåíèÿ (ïîäâèæíîñòè) áåëêîâ â ýëåêòðè÷åñêîì ïîëå, êîòîðàÿ Ðèñ. 1.5. Ãåëü-õðîìàòîãðàôèÿ íà êîëîíêå ñ ñåôàäåêñîì (ñõåìà). Áîëüøèå ñâåòëûå êðóæêè ñ êðåñòèêàìè - çåðíà ñåôàäåêñà; ìàëûå ÷åðíûå è êðàñíûå êðóæêè è òðåóãîëüíèêè - áåëêè ñ ðàçëè÷íîé ìîëåêóëÿðíîé ìàññîé; À - êîëîíêà â íà÷àëå ðàáîòû; Á, Â, Ã - êîëîíêà â ðàçëè÷íûå ïåðèîäû âðåìåíè. Íà ãðàôèêå ýëþöèè ÷åòêî âèäíî ðàçäåëåíèå áåëêîâûõ êîìïîíåíòîâ. îïðåäåëÿåòñÿ âåëè÷èíîé çàðÿäà áåëêà ïðè îïðåäåëåííûõ çíà÷åíèÿõ ðÍ è èîííîé ñèëû ðàñòâîðà. Â ïîñëåäíåå âðåìÿ áîëåå øèðîêîå ðàñïðîñòðàíåíèå ïîëó÷èëè ìåòîäû çîíàëüíîãî ýëåêòðîôîðåçà áåëêîâ íà ðàçëè÷íûõ íîñèòåëÿõ, â ÷àñòíîñòè íà òâåðäûõ ïîääåðæèâàþùèõ ñðåäàõ: ãåëÿõ êðàõìàëà è ïîëèàêðèëàìèäà, öåëëþëîçå. Ïðåèìóùåñòâà èõ ïî ñðàâíåíèþ ñ ìåòîäîì ñâîáîäíîãî ýëåêòðîôîðåçà ñîñòîÿò â òîì, ÷òî èñêëþ÷àåòñÿ ðàçìûâàíèå ãðàíèöû áåëîê-ðàñòâîðèòåëü â ðåçóëüòàòå äèôôóçèè è êîíâåêöèè, íå òðåáóåòñÿ íàëàæèâàíèÿ ñëîæíîé àïïàðàòóðû äëÿ îïðåäåëåíèÿ ïîëîæåíèÿ ãðàíèöû, à äëÿ àíàëèçà íåîáõîäèìî íåáîëüøîå êîëè÷åñòâî áåëêà (ïîäðîáíî ýòè ìåòîäû è ñîîòâåòñòâóþùàÿ àïïàðàòóðà ðàññìàòðèâàþòñÿ â ïðàêòè÷å-ñêèõ ðóêîâîäñòâàõ ïî áèîõèìèè). Îäíèì èç íàèáîëåå ðàñïðîñòðàíåííûõ ìåòîäîâ ôðàêöèîíèðîâàíèÿ áåëêîâ (êàê è ìåòîäîâ îöåíêè ãîìîãåííîñòè) ÿâëÿåòñÿ äèñê-ýëåêòðîôîðåç (îò àíãë. ØçñîïÈïèîèç - ïðåðûâèñòûé, ïåðåìåæàþùèéñÿ) â ïîëèàêðèëàìèäíîì ãåëå, ïðè êîòîðîì èñïîëüçóþò ïàðû áóôåðíûõ ðàñòâîðîâ ñ ðàçëè÷íûìè çíà÷åíèÿìè ðÍ è ðàçíîé ñòåïåíè ïîðèñòîñòè ãåëü. Ñëåäóåò îòìåòèòü âûñîêóþ ðàçðåøàþùóþ ñïîñîáíîñòü ãåëü-ýëåêòðîôîðåçà. Åñëè ïðè ýëåêòðîôîðåçå áåëêîâ ñûâîðîòêè êðîâè ÷åëîâåêà íà áóìàãå îòêðûâàþòñÿ âñåãî 6 ôðàêöèé, òî ïðè ýëåêòðîôîðåçå â êðàõìàëüíîì ãåëå - 10, à â ïîëèàêðèëàìèäíîì ãåëå - äî 18 ðàçíûõ áåëêîâûõ ôðàêöèé. Äëÿ âûÿâëåíèÿ áåëêîâ ïðè ýëåêòðîôîðåçå â ãåëÿõ èõ îáðàáàòûâàþò îäíèì èç ñëåäóþùèõ êðàñèòåëåé: áðîìôåíîëîâûì ñèíèì, àìèäî ÷åðíûì 10Â, êèñëîòíûì ñèíèì 83, êóìàññè áðèëëèàíòîâûì ãîëóáûì Ê-250 è äð. Èíòåíñèâíîñòü îêðàñêè è ñîîòâåòñòâåííî îòíîñèòåëüíîå ñîäåðæàíèå êàæäîé áåëêîâîé ôðàêöèè îáû÷íî îïðåäåëÿþò äåíñèòîìåòðè÷åñêè ïóòåì ïðÿìîãî ñêàíèðîâàíèÿ íà äåíñèòîìåòðå. Â ïîñëåäíèå ãîäû ñòàëè ïðèìåíÿòü ìåòîäû ýëåêòðîôîðåçà áåëêîâ ñ ãðàäèåíòîì êîíöåíòðàöèè ãåëÿ, ÷òî çíà÷èòåëüíî ïîâûøàåò ðàçðåøàþùóþ ñïîñîáíîñòü, îñîáåííî ïðè ôðàêöèîíèðîâàíèè áåëêîâ ñ âûñîêîé ìîëåêóëÿðíîé ìàññîé, ïðåâûøàþùåé 50000— 100000. Âåñüìà ïåðñïåêòèâíûìè ìåòîäàìè ðàçäåëåíèÿ áåëêîâ (êàê è îïðåäåëåíèÿ ðÿäà ôèçèêî-õèìè÷åñêèõ ñâîéñòâ) îêàçàëèñü ðàçíûå âàðèàíòû ìåòîäà èçîýëåêòðè÷åñêîãî ôîêóñèðîâàíèÿ - èçîòàõîôîðåçà, îñíîâàííûå íà ïðîâåäåíèè ýëåêòðîôîðåçà â ïîääåðæèâàþùèõ ñðåäàõ (íà êîëîíêå èëè â òîíêîì ñëîå) ñ ãðàäèåíòîì ðÍ. Òî÷íîå ìåñòîïîëîæåíèå íà êîëîíêå êàæäîãî áåëêà èç ñìåñè îïðåäåëÿåòñÿ çíà÷åíèåì åãî èçîýëåêòðè÷åñêîé òî÷êè, ò.å. ñîñòîÿíèåì, ïðè êîòîðîì ñóììàðíûé ýëåêòðè÷åñêèé çàðÿä áåëêîâîé ÷àñòèöû ïðè äàííîì çíà÷åíèè ðÍ ðàâåí íóëþ. Ïðè èñïîëüçîâàíèè ìåòîäà èçîýëåêòðè÷åñêîãî ôîêóñèðîâàíèÿ ïðèìåíÿþò ñìåñè ñèíòåòè÷åñêèõ ïîëèàìèíîïîëèêàðáîíîâûõ êèñëîò (àìôîëèíû) äëÿ ñîçäàíèÿ ãðàäèåíòà ðÍ â äèàïàçîíå îò 3,0 äî 10,0. Â ïîñëåäíèå ãîäû øèðîêîå ðàñïðîñòðàíåíèå äëÿ ôðàêöèîíèðîâàíèÿ áåëêîâ ïîëó÷èëè ðàçëè÷íûå ñî÷åòàíèÿ èçîýëåêòðîôîêóñèðîâàíèÿ è äèñê- ýëåêòðîôîðåçà â ïîëèàêðèëàìèäíîì ãåëå—ìåòîäû äâóõìåðíîãî ýëåêòðîôîðåçà, êîòîðûå ïîçîëÿþò ïàðàëëåëüíî àíàëèçèðîâàòü ñîòíè è äàæå òûñÿ÷è áåëêîâûõ ôðàêöèé. Î÷èñòêà áåëêîâ îò íèçêîìîëåêóëÿðíûõ ïðèìåñåé Ïðèìåíåíèå â îïðåäåëåííîé ïîñëåäîâàòåëüíîñòè ðÿäà ïåðå÷èñëåííûõ ìåòîäîâ ïîçâîëÿåò ïîëó÷èòü áåëîê â î÷èùåííîì ñîñòîÿíèè, íå ëèøåííûé, îäíàêî, íåêîòîðûõ ïðèìåñåé ñîëåé. Äëÿ ïîëíîãî îñâîáîæäåíèÿ áåëêîâ îò íèçêîìîëåêóëÿðíûõ ïðèìåñåé â íàñòîÿùåå âðåìÿ èñïîëüçóþò ìåòîäû äèàëèçà, ãåëüõðîìàòîãðàôèè, êðèñòàëëèçàöèè, óëüòðàôèëüòðàöèè. Ïðè äèàëèçå ïðèìåíÿþò ïîëóïðîíèöàåìûå ìåìáðàíû (öåëëîôàí, êîëëîäèéíàÿ ïëåíêà), äèàìåòð ïîð êîòîðûõ âàðüèðóåò â øèðîêèõ ïðåäåëàõ. Áåëêè, êàê ïðàâèëî, íå äèôôóíäèðóþò ÷åðåç òàêóþ ìåìáðàíó, â òî âðåìÿ êàê íèçêîìîëåêóëÿðíûå âåùåñòâà ëåãêî ïðîíèêàþò ÷åðåç íåå â îêðóæàþùóþ ñðåäó. Ìåòîä êðèñòàëëèçàöèè áåëêîâ îñíîâàí íà äîñòèæåíèè êðèòè÷åñêîé òî÷êè íà÷àëà îñàæäåíèÿ áåëêà èç ðàñòâîðà ñóëüôàòà àììîíèÿ ïðè ìåä-ëåííîì ïîâûøåíèè òåìïåðàòóðû. Óæå ïîëó÷åíû ñîòíè êðèñòàëëè÷åñêèõ áåëêîâ . Îäíàêî íå âñÿêèé êðèñòàëëè÷åñêèé áåëîê ÿâëÿåòñÿ ãîìîãåííûì, ïîñêîëüêó ïðè îäíîé è òîé æå êîíöåíòðàöèè ðàñòâîðà ñóëüôàòà àììîíèÿ ìîãóò êðèñòàëëèçîâàòüñÿ áëèçêèå ïî ðàçìåðàì è ìàññå ðàçíûå áåëêè. Íàèëó÷øèå ðåçóëüòàòû ïðè îñâîáîæäåíèè áåëêîâ îò íèçêîìîëåêóëÿðíûõ ïðèìåñåé ïîëó÷àþò ñ ïîìîùüþ ãåëüõðîìàòîãðàôèè è óëüòðàôèëüòðàöèè. Ïîñëåäíÿÿ îñíîâàíà íà ïðîäàâëèâàíèè ðàñòâîðîâ áåëêà ÷åðåç ñïåöèàëüíûå ìåìáðàíû, çàäåðæèâàþùèå áåëêîâûå ìîëåêóëû, ÷òî ïîçâîëÿåò íå òîëüêî îñâîáîäèòü áåëêîâûå ðàñòâîðû îò íèçêîìîëåêóëÿðíûõ ïðèìåñåé, íî è êîíöåíòðèðîâàòü èõ. Îïðåäåëåíèå ãîìîãåííîñòè áåëêîâ Íà çàêëþ÷èòåëüíîì ýòàïå âûäåëåíèÿ è î÷èñòêè áåëêîâ èññëåäîâàòåëÿ âñåãäà èíòåðåñóåò âîïðîñ î ãîìîãåííîñòè ïîëó÷åííîãî áåëêà. Íåëüçÿ îöåíèâàòü ãîìîãåííîñòü èíäèâèäóàëüíîãî áåëêà òîëüêî ïî îäíîìó êàêîìó- ëèáî ôèçèêî-õèìè÷åñêîìó ïîêàçàòåëþ. Äëÿ ýòîãî ïîëüçóþòñÿ ðàçíûìè êðèòåðèÿìè. Èç îãðîìíîãî ÷èñëà õðîìàòîãðàôè÷åñêèõ, ýëåêòðîôîðåòè÷åñêèõ, õèìè÷åñêèõ, ðàäèî- è èììóíîõèìè÷åñêèõ, áèîëîãè÷åñêèõ è ãðàâèòàöèîííûõ ìåòîäîâ íàèáîëåå äîñòîâåðíûå ðåçóëüòàòû ïðè îïðåäåëåíèè ãîìîãåííîñòè áåëêà äàþò óëüòðàöåíòðèôóãèðîâàíèå â ãðàäèåíòå ïëîòíîñòè ñàõàðîçû èëè õëîðèäà öåçèÿ, äèñê-ýëåêòðîôîðåç â ïîëèàêðèëàìèäíîì ãåëå, èçîýëåêòðè÷åñêîå ôîêóñèðîâàíèå, èììóíîõèìè÷åñêèå ìåòîäû è îïðåäåëåíèå ðàñòâîðèìîñòè áåëêà. Äåéñòâèòåëüíî, åñëè ïðè ãåëü-ýëåêòðîôîðåçå áåëîê äâèæåòñÿ â âèäå îäíîé óçêîé ïîëîñû è â ýòîé çîíå ñîñðåäîòî÷åíà åãî áèîëîãè÷åñêàÿ àêòèâíîñòü (ôåðìåíòàòèâíàÿ, ãîðìîíàëüíàÿ, òîêñè÷åñêàÿ è ò.ä.), òî ýòè äàííûå ñ áîëüøîé äîëåé âåðîÿòíîñòè ìîãóò ñâèäåòåëüñòâîâàòü îá îäíîðîäíîñòè èññëåäóåìîãî áåëêà. Â îñíîâå èììóíîõèìè÷åñêîãî ìåòîäà êîíòðîëÿ ãîìîãåííîñòè èññëåäóåìîãî áåëêà ëåæèò ðåàêöèÿ ïðåöèïèòàöèè åãî ñ ñîîòâåòñòâóþùåé àíòèñûâîðîòêîé, ïîëó÷åííîé îò èììóíèçèðîâàííûõ ýòèì áåëêîì æèâîòíûõ. Äëÿ ñòðîãîãî äîêàçàòåëüñòâà ãîìîãåííîñòè áåëêà òðåáóåòñÿ îäíîâðåìåííîå èñïîëüçîâàíèå íåñêîëüêèõ ìåòîäîâ. Íå ïîòåðÿë ñâîåãî çíà÷åíèÿ è ìåòîä êðèñòàëëèçàöèè áåëêîâ ñ èñïîëüçîâàíèåì ñóëüôàòà àììîíèÿ, à òàêæå ìåòîä îïðåäåëåíèÿ ðàñòâîðèìîñòè áåëêà. Ïîñëåäíèé, ïðåäëîæåííûé åùå Ä. Íîðòðîïîì , îñíîâàí íà ïðàâèëå ôàç Ãèááñà, ñîãëàñíî êîòîðîìó ðàñòâîðèìîñòü ÷èñòîãî âåùåñòâà ïðè äàííûõ óñëîâèÿõ îïûòà çàâèñèò òîëüêî îò òåìïåðàòóðû, íî íå çàâèñèò îò êîëè÷åñòâà âåùåñòâà, íàõîäÿùåãîñÿ â òâåðäîé ôàçå. Ìåòîä ìîæåò áûòü âûïîëíåí ñðàâíèòåëüíî ëåãêî è áûñòðî â ìèêðîìàñøòàáàõ. Îáû÷íî îïðåäåëÿþò ðàñòâîðèìîñòü óâåëè÷èâàþùåãîñÿ êîëè÷åñòâà èññëåäóåìîãî áåëêà ïðè ïîñòîÿííîì êîëè÷åñòâå ðàñòâîðèòåëÿ. ÀÌÈÍÎÊÈÑËÎÒÍÛÉ ÑÎÑÒÀÂ ÁÅËÊÎÂ Íåñìîòðÿ íà òî ÷òî ïåðâàÿ àìèíîêèñëîòà—ãëèöèí—áûëà âûäåëåíà À. Áðà- êîííî åùå â 1820 ã. èç êèñëîòíîãî ãèäðîëèçàòà æåëàòèíà, ïîëíûé àìèíîêèñëîòíûé ñîñòàâ áåëêîâ áûë ðàñøèôðîâàí òîëüêî ê 30-ì ãîäàì XX â. Áîëüøàÿ çàñëóãà â ýòîì ïðèíàäëåæèò ðàáîòàì Í.Í. Ëþáàâèíà, êîòîðûé â 1871 ã. óñòàíîâèë, ÷òî ïîä äåéñòâèåì ôåðìåíòîâ ïèùåâàðèòåëüíûõ ñîêîâ áåëêè ðàñùåïëÿþòñÿ íà àìèíîêèñëîòû. Áûëè ñäåëàíû äâà âàæíûõ âûâîäà: 1) â ñîñòàâ áåëêîâ âõîäÿò àìèíîêèñëîòû; 2) ìåòîäàìè ãèäðîëèçà ìîæåò áûòü èçó÷åí õèìè÷åñêèé, â ÷àñòíîñòè àìíîêèñëîòíûé, ñîñòàâ áåëêîâ. Äëÿ èçó÷åíèÿ àìèíîêèñëîòíîãî ñîñòàâà áåëêîâ ïîëüçóþòñÿ ñî÷åòàíèåì êèñëîòíîãî (ÍÑ1), ùåëî÷íîãî [Âà(ÎÍ)2] è, ðåæå, ôåðìåíòàòèâíîãî ãèäðîëèçà èëè îäíèì èç íèõ. Óñòàíîâëåíî, ÷òî ïðè ãèäðîëèçå ÷èñòîãî áåëêà, íå ñîäåðæàùåãî ïðèìåñåé, îñâîáîæäàþòñÿ 20 ðàçëè÷íûõ à-àìèíîêèñëîò. Âñå äðóãèå îòêðûòûå â òêàíÿõ æèâîòíûõ, ðàñòåíèé è ìèêðîîðãàíèçìîâ àìèíîêèñëîòû (áîëåå 300) ñóùåñòâóþò â ïðèðîäå â ñâîáîäíîì ñîñòîÿíèè ëèáî â âèäå êîðîòêèõ ïåïòèäîâ èëè êîìïëåêñîâ ñ äðóãèìè îðãàíè÷åñêèìè âåùåñòâàìè. à-Àìèíîêèñëîòû ïðåäñòàâëÿþò ñîáîé ïðîèçâîäíûå êàðáîíîâûõ êèñëîò, ó êîòîðûõ îäèí âîäîðîäíûé àòîì, ó à-óãëåðîäà, çàìåùåí íà àìèíîãðóïïó (—ÊÈ2), íàïðèìåð: Ð—ÑÍ2—ÑÍ2—ÑÎÎÈ Ê—ÑÍ—ÑÎÎÍ NN3 à-Àìèíîêèñëîòà Ñëåäóåò ïîä÷åðêíóòü, ÷òî âñå àìèíîêèñëîòû, âõîäÿùèå â ñîñòàâ ïðèðîäíûõ áåëêîâ, ÿâëÿþòñÿ à-àìèíîêèñëîòàìè, õîòÿ àìèíîãðóïïà â ñâîáîäíûõ àìèíîêàðáîíîâûõ êèñëîòàõ ìîæåò íàõîäèòüñÿ, êàê óâèäèì íèæå, â Ð-, ó-, áè å-ïîëîæåíèÿõ. Êëàññèôèêàöèÿ àìèíîêèñëîò Âñå âñòðå÷àþùèåñÿ â ïðèðîäå àìèíîêèñëîòû îáëàäàþò îáùèì ñâîéñòâîì - àìôîòåðíîñòüþ (îò ãðå÷. àøðÜî1åãî§ - äâóñòîðîííèé), ò.å. êàæäàÿ àìèíîêèñëîòà ñîäåðæèò êàê ìèíèìóì îäíó êèñëîòíóþ è îäíó îñíîâíóþ ãðóïïû. Îáùèé òèï ñòðîåíèÿ à-àìèíîêèñëîò ìîæåò áûòü ïðåäñòàâëåí â ñëåäóþùåì âèäå: È"—Ñ—N Í2 Êàê âèäíî èç îáùåé ôîðìóëû, àìèíîêèñëîòû áóäóò îòëè÷àòüñÿ äðóã îò äðóãà õèìè÷åñêîé ïðèðîäîé ðàäèêàëà Ê, ïðåäñòàâëÿþùåãî ãðóïïó àòîìîâ â ìîëåêóëå àìèíîêèñëîòû, ñâÿçàííóþ ñ à-óãëåðîäíûì àòîìîì è íå ó÷àñòâóþùóþ â îáðàçîâàíèè ïåïòèäíîé ñâÿçè ïðè ñèíòåçå áåëêà. Ïî÷òè âñå à-àìèíî- è à-êàðáîêñèëüíûå ãðóïïû ó÷àñòâóþò â îáðàçîâàíèè ïåïòèäíûõ ñâÿçåé áåëêîâîé ìîëåêóëû, òåðÿÿ ïðè ýòîì ñâîè ñïåöèôè÷åñêèå äëÿ ñâîáîäíûõ àìèíîêèñëîò êèñëîòíî-îñíîâíûå ñâîéñòâà. Ïîýòîìó âñå ðàçíîîáðàçèå îñîáåííîñòåé ñòðóêòóðû è ôóíêöèè áåëêîâûõ ìîëåêóë ñâÿçàíî ñ õèìè÷åñêîé ïðèðîäîé è ôèçèêî-õèìè÷åñêèìè ñâîéñòâàìè ðàäèêàëîâ àìèíîêèñëîò. Èìåííî áëàãîäàðÿ èì áåëêè íàäåëåíû ðÿäîì óíèêàëüíûõ ôóíêöèé, íå ñâîéñòâåííûõ äðóãèì áèîïîëèìåðàì, è îáëàäàþò õèìè÷åñêîé èíäèâèäóàëüíîñòüþ. Êëàññèôèêàöèÿ àìèíîêèñëîò ðàçðàáîòàíà íà îñíîâå õèìè÷åñêîãî ñòðîåíèÿ ðàäèêàëîâ, õîòÿ áûëè ïðåäëîæåíû è äðóãèå ïðèíöèïû. Ðàçëè÷àþò àðîìàòè÷åñêèå è àëèôàòè÷åñêèå àìèíîêèñëîòû, à òàêæå àìèíîêèñëîòû, ñîäåðæàùèå ñåðó èëè ãèäðîêñèëüíûå ãðóïïû. ×àñòî êëàññèôèêàöèÿ îñíîâàíà íà ïðèðîäå çàðÿäà àìèíîêèñëîòû. Åñëè ðàäèêàë íåéòðàëüíûé (òàêèå àìèíîêèñëîòû ñîäåðæàò òîëüêî îäíó àìèíî- è îäíó êàðáîêñèëüíóþ ãðóïïû), òî îíè íàçûâàþòñÿ íåéòðàëüíûìè àìèíîêèñëîòàìè. Åñëè àìèíîêèñëîòà ñîäåðæèò èçáûòîê àìèíî- èëè êàðáîêñèëüíûõ ãðóïï, òî îíà íàçûâàåòñÿ ñîîòâåòñòâåííî îñíîâíîé èëè êèñëîé àìèíîêèñëîòîé. Ñîâðåìåííàÿ ðàöèîíàëüíàÿ êëàññèôèêàöèÿ àìèíîêèñëîò îñíîâàíà íà ïîëÿðíîñòè ðàäèêàëîâ (Ê-ãðóïï), ò.å. ñïîñîáíîñòè èõ ê âçàèìîäåéñòâèþ ñ âîäîé ïðè ôèçèîëîãè÷åñêèõ çíà÷åíèÿõ ðÍ (áëèçêèõ ê ðÍ 7,0). Ðàçëè÷àþò 5 êëàññîâ àìèíîêèñëîò, ñîäåðæàùèõ ñëåäóþùèå ðàäèêàëû: 1) íåïîëÿðíûå (ãèäðîôîáíûå); 2) ïîëÿðíûå (ãèäðîôèëüíûå); 3) àðîìàòè÷åñêèå (áîëüøåé ÷àñòüþ íåïîëÿðíûå); 4) îòðèöàòåëüíî çàðÿæåííûå è 5) ïîëîæèòåëüíî çàðÿæåííûå. Â ïðåäñòàâëåííîé êëàññèôèêàöèè àìèíîêèñëîò (òàáë. 1.3) ïðèâåäåíû íàèìåíîâàíèÿ, ñîêðàùåííûå àíãëèéñêèå è ðóññêèå îáîçíà÷åíèÿ è îäíîáóêâåííûå ñèìâîëû àìèíîêèñëîò, ïðèíÿòûå â îòå÷åñòâåííîé è èíîñòðàííîé ëèòåðàòóðå, à òàêæå çíà÷åíèÿ èçîýëåêòðè÷åñêîé òî÷êè (ð1) è ìîëåêóëÿðíîé ìàññû (Ì). Îòäåëüíî äàþòñÿ ñòðóêòóðíûå ôîðìóëû âñåõ 20 àìèíîêèñëîò áåëêîâîé ìîëåêóëû. Àìèíîêèñëîòû Ïðèíÿòûå ñîêðàùåííûå îáîçíà÷åíèÿ è îäíîáóêâåííûå ñèìâîëû Ì/ð! Ñðåäíåå ñîäåðæàíèå â áåëêàõ, % àíãë. ñèìâîë ðóññê. I. Íåïîëÿðíûå Ê-ãðóïïû Ãëèöèí Ñ1ó î Ãëè 75/5,97 7,5 Àëàíèí À1à À Àëà 89/6,02 9,0 Âàëèí Óà1 Ó Âàë 117/5,97 6,9 Ëåéöèí Ãåè ü Ëåé 131/5,97 7,5 Èçîëåéöèí Íå I Èëå 131/5,97 4,6 Ïðîëèí Ðãî Ð Ïðî 115/6,10 4,6 II. Ïîëÿðíûå, íåçàðÿæåííûå Ê-ãðóïïû Ñåðèí 8åã 8 Ñåð 105/5,68 7,1 Òðåîíèí ÒÜã Ò Òðå 119/6,53 6,0 Öèñòåèí Ñóê Ñ Öèñ 121/5,02 2,8 Ìåòèîíèí Ìå! Ì Ìåò 149/5,75 1,7 Àñïàðàãèí Àêï N Àñí 132/5,41 4,4 Ãëóòàìèí Ñ1ï 0 Ãëí 146/5,65 3,9 III. Àðîìàòè÷åñêèå Ê- ãðóïïû Ôåíèëàëàíèí ÐÜå ð Ôåí 165/5,98 3,5 Òèðîçèí Òóã V Òèð 181/5,65 3,5 Òðèïòîôàí Òãð ¹ Òðï 204/5,88 1,1 ÃÓ. Îòðèöàòåëüíî çàðÿ-æåííûå Ê-ãðóïïû Àñïàðàãèíîâàÿ Àêð Á Àñï 133/2,97 5,5 êèñëîòà Ãëóòàìèíîâàÿ Ñ1è Å Ãëó 147/3,22 6,2 êèñëîòà Ó. Ïîëîæèòåëüíî çàðÿ-æåííûå Ê-ãðóïïû Ëèçèí Ãóê Ê Ëèç 146/9,74 7,0 Àðãèíèí Àãä Ê Àðã 174/10,76 4,7 Ãèñòèäèí Øê Í Ãèñ 155/7,59 2,1 Íåïîëÿðíûå Ï-ãðóïïû 1_-ãëóòàìèí î=ñ-ñíãÒñí-ñîîí 1 I ¦ I I Îòðèöàòåëüíî çàðÿæåííûå Ð-ãðóïïû ÍÎÎÎÑÍã-ÑÍ2òÑÍ-ÑÎÎÍ \| ! ìí, I ÍÎÎÑ-ÑÍ21ÑÍ-ÑÎÎÍ I I ÛÍã I 1_-àñïàðàãèíîâàÿ êèñëîòà Ïîëîæèòåëüíî çàðÿæåííûå Ð-ãðóïïû ( I 1 ÑÍ,-ÑÍÝ-ÑÍ,-ÑÍ5+ÑÍ~ÑÎÎÍ ! /=ò-ÑÍã-ÑÍ-ÑÎÎÍ ] Ü1Í2 ^ 1 ÍÌ÷,Ì I 1_-ãèñòèäèí NN , Ñ-ÌÍ-ÑÍ,-ÑÍ,-ÑÍ,+ÑÍ-ÑÎÎÍ ^ . !Ã4Í2 ¦ 1_-àðãèíèí I Àðîìàòè÷åñêèå Ð-ãðóïïû 1_-òðèïòîôàí Ïåðå÷èñëåííûå àìèíîêèñëîòû ïðèñóòñòâóþò â ðàçíûõ êîëè÷åñòâåííûõ ñîîòíîøåíèÿõ è ïîñëåäîâàòåëüíîñòÿõ â òûñÿ÷àõ áåëêîâ, õîòÿ îòäåëüíûå èíäèâèäóàëüíûå áåëêè íå ñîäåðæàò ïîëíîãî íàáîðà âñåõ ýòèõ àìèíîêèñëîò. Ïîìèìî íàëè÷èÿ â áîëüøèíñòâå ïðèðîäíûõ áåëêîâ 20 àìèíîêèñëîò, â íåêîòîðûõ áåëêàõ îáíàðóæåíû ïðîèçâîäíûå àìèíîêèñëîò : îêñèïðîëèí, îêñèëèçèí, äèéîäòèðîçèí, ôîñôîñåðèí è ôîñôîòðåîíèí (ïîñëåäíèå äâå àìèíîêèñëîòû ïðåäñòàâëåíû â ãëàâå 2): ÑÍý-ÑÍ-(ÑÍ2)2-ÑÍ-ÑÎÎÍ è^>í 1\|í2 Îêñèëèçèí Ïåðâûå äâå àìèíîêèñëîòû ñîäåðæàòñÿ â áåëêå ñîåäèíèòåëüíîé òêàíè - êîëëàãåíå, à äèéîäòèðîçèí ÿâëÿåòñÿ îñíîâîé ñòðóêòóðû ãîðìîíîâ ùèòîâèäíîé æåëåçû. Â ìûøå÷íîì áåëêå ìèîçèíå îáíàðóæåí òàêæå å-Ì-ìåòèëëèçèí; â ñîñòàâ ïðîòðîìáèíà (áåëîê ñâåðòûâàíèÿ êðîâè) âõîäèò ó-êàðáîêñèãëóòà- ìèíîâàÿ êèñëîòà, à â ãëóòàòèîíïåðîêñèäàçå îòêðûò ñåëåíîöèñòåèí, â êîòîðîì ÎÍ-ãðóïïà ñåðèíà çàìåíåíà íà ñåëåí (8å): 7 Ïîìèìî óêàçàííûõ, ðÿä à-àìèíîêèñëîò âûïîëíÿåò âàæíûå ôóíêöèè â îáìåíå âåùåñòâ, õîòÿ è íå âõîäèò â ñîñòàâ áåëêîâ, â ÷àñòíîñòè îðíèòèí, öèòðóëëèí, ãîìîñåðèí, ãîìîöèñòåèí, öèñòåèíñóëüôèíîâàÿ êèñëîòà, äèîêñè- ôåíèëàëàíèí è äð. Îáùèå ñâîéñòâà àìèíîêèñëîò Êèñëîòíî-îñíîâíûå ñâîéñòâà. Ýòè ñâîéñòâà àìèíîêèñëîò îïðåäåëÿþò ìíîãèå ôèçèêî-õèìè÷åñêèå è áèîëîãè÷åñêèå ñâîéñòâà áåëêîâ. Íà ýòèõ ñâîéñòâàõ îñíîâàíû, êðîìå òîãî, ïî÷òè âñå ìåòîäû âûäåëåíèÿ è èäåíòèôèêàöèè àìèíîêèñëîò. Àìèíîêèñëîòû ëåãêî ðàñòâîðèìû â âîäå. Îíè êðèñòàëëèçóþòñÿ èç íåéòðàëüíûõ âîäíûõ ðàñòâîðîâ â ôîðìå áèïîëÿðíûõ (àìôîòåð- íûõ) èîíîâ (öâèòòåðèîíîâ), à íå â âèäå íåäèññîöèèðîâàííûõ ìîëåêóë (ïîñëåäíþþ ñòðóêòóðó ïðèâîäÿò äëÿ óäîáñòâà ïðåäñòàâëåíèÿ, îäíàêî âñå àìèíîêèñëîòû ïðè ôèçèîëîãè÷åñêèõ çíà÷åíèÿõ ðÍ èìåþò ñòðóêòóðó öâèòòå- ðèîíà). Í—Ñ-ÊÍ2 I ÑÎÎÍ Ïðè ðàñòâîðåíèè â âîäå êðèñòàëëè÷åñêàÿ àìèíîêèñëîòà, íàïðèìåð àëàíèí, ìîæåò ðåàãèðîâàòü èëè êàê êèñëîòà (äîíàòîð ïðîòîíà): +1×Í3ÑÍ(ÑÍ3)ÑÎÎ-<=> Í+ + 1×Í2ÑÍ(ÑÍ3)ÑÎÎ-, èëè êàê îñíîâàíèå (àêöåïòîð ïðîòîíà): +ÛÍ3ÑÍ(ÑÍ3)ÑÎÎ-+Í+<=>+ÛÍ3ÑÍ(ÑÍ3)ÑÎÎÍ. Åñëè ðàäèêàëû àìèíîêèñëîò íåéòðàëüíûå, òî îíè ïî÷òè íå îêàçûâàþò âëèÿíèÿ íà äèññîöèàöèþ à-êàðáîêñèëüíîé ãðóïïû èëè à-àìèíîãðóïïû, è âåëè÷èíû ðÊ (îòðèöàòåëüíûé ëîãàðèôì êîíñòàíòû äèññîöèàöèè) îñòàþòñÿ îòíîñèòåëüíî ïîñòîÿííûìè. Âñëåäñòâèå ýòîãî êðèâûå äèññîöèàöèè ïî÷òè âñåõ íåéòðàëüíûõ àìèíîêèñëîò íàêëàäûâàþòñÿ äðóã íà äðóãà è ìîãóò áûòü ðàññìîòðåíû íà ïðèìåðå àëàíèíà. Åñëè ê ðàñòâîðó àëàíèíà (íàïðèìåð, 1 Ì) â âîäå ïîñòåïåííî ïðèáàâëÿòü ñèëüíóþ êèñëîòó (0,1 Ì ðàñòâîð ÍÑ1) èëè ñèëüíóþ ùåëî÷ü (0,1 Ì ðàñòâîð ¹ÎÍ), òî ïîëó÷èì êðèâóþ òèòðîâà-íèÿ àëàíèíà, òèïè÷íóþ äëÿ âñåõ íåéòðàëüíûõ àìèíîêèñëîò (ðèñ. 1.6). Êàæóùèåñÿ âåëè÷èíû ðÊ' äëÿ à-êàðáîêñèëüíîé ãðóïïû è à-àìèíîãðóïï (ò.å. çíà÷åíèÿ ðÍ, ïðè êîòîðûõ ýòè ãðóïïû â ñðåäíåì íàïîëîâèíó äèññîöèèðîâàíû) äîâîëüíî ñèëüíî ðàçëè÷àþòñÿ, ñîñòàâëÿÿ ðÊ1 = 2,34 è ðÊ2 = 9,69. Ïðè íèçêîì çíà÷åíèè ðÍ (íèæå ðÊ1') ïî÷òè âñå ìîëåêóëû àëàíèíà ÿâëÿþòñÿ ïîëíîñòüþ ïðîòîíèðîâàííûìè è íåñóò ïîëîæèòåëüíûé çàðÿä. Äðóãèìè ñëîâàìè, ïðè âûñîêîé êîíöåíòðàöèè âîäîðîäíûõ èîíîâ â ðàñòâîðå òåíäåíöèÿ ê äèññîöèàöèè âîäîðîäà èç ñòðóêòóðû àëàíèíà îêàçûâàåòñÿ íåçíà÷èòåëüíîé. Èç êðèâîé òèòðîâàíèÿ âèäíî, ÷òî òî÷êà ïåðåõîäà ìåæäó âåòâÿìè êðèâîé ðàñïîëàãàåòñÿ ïðè ðÍ 6,02. Ýòî îçíà÷àåò, ÷òî ïðè äàííîì çíà÷åíèè ðÍ ñóììàðíûé (èëè ñðåäíèé) ýëåêòðè÷åñêèé çàðÿä ìîëåêóëû àëàíèíà ðàâåí íóëþ è îíà íå ïåðåìåùàåòñÿ â ýëåêòðè÷åñêîì ïîëå íè ê àíîäó, íè ê êàòîäó (èçîýëåêòðè÷åñêîå ñîñòîÿíèå). Òàêîå çíà÷åíèå ðÍ ïîëó÷èëî íàçâàíèå èçî- ýëåêòðè÷åñêîé òî÷êè è îáîçíà÷àåòñÿ ð1. Èçîýëåêòðè÷åñêàÿ òî÷êà àìèíîêèñëîò, íå ñîäåðæàùèõ äîïîëíèòåëüíûõ N4.,- èëè ÑÎÎÍ-ãðóïï, ïðåäñòàâëÿåò ñîáîé ñðåäíåå àðèôìåòè÷åñêîå ìåæäó äâóìÿ çíà÷åíèÿìè ðÊ': ðÊ'ñîîí + ðê'ìí, ð\| = 2 ’ ñîîòâåòñòâåííî äëÿ àëàíèíà 2,34 + 9,69 „ ð\| = 2 6’ Èçîýëåêòðè÷åñêàÿ òî÷êà ðÿäà äðóãèõ àìèíîêèñëîò, ñîäåðæàùèõ äîïîëíèòåëüíûå êèñëîòíûå èëè îñíîâíûå ãðóïïû (àñïàðàãèíîâàÿ è ãëóòàìèíîâàÿ êèñëîòû, ëèçèí, àðãèíèí, òèðîçèí è äð.), çàâèñèò, êðîìå òîãî, îò êèñëîòíîñòè èëè îñíîâíîñòè ðàäèêàëîâ ýòèõ àìèíîêèñëîò. Äëÿ ëèçèíà, íàïðèìåð, ð1 äîëæíà âû÷èñëÿòüñÿ èç ïîëóñóììû çíà÷åíèé ðÊ' äëÿ à- è å-ÊÍ2-ãðóïï. Òàêèì îáðàçîì, â èíòåðâàëå ðÍ îò 4,0 äî 9,0 ïî÷òè âñå àìèíîêèñëîòû ñóùåñòâóþò ïðåèìóùåñòâåííî â ôîðìå öâèòòåðèîíîâ ñ ïðîòîíèðîâàííîé àìèíîãðóïïîé è äèññîöèèðîâàííîé êàðáîêñèëüíîé ãðóïïîé. Ñëåäóåò îòìå- Ðèñ. 1.6. Êðèâûå, ïîëó÷åííûå ïðè òèòðîâàíèè 0,1 Ì ðàñòâîðà àëàíèíà 0,1 Ì ðàñòâîðîì ÍÑ1 (à) è 0,1 Ì ðàñòâîðîì ¹ÎÍ (á). òèòü, ÷òî ïðè ôèçèîëîãè÷åñêèõ çíà÷åíèÿõ ðÍ òêàíåé è êðîâè (7,1 è 7,4 ñîîòâåòñòâåííî) àìèíîêèñëîòû (çà èñëþ÷åíèåì ãèñòèäèíà) íå îáëàäàþò èçìåðèìîé áóôåðíîé åìêîñòüþ. Ýòó ñïîñîáíîñòü îíè ïðèîáðåòàþò òîëüêî ïðè çíà÷åíèÿõ ðÍ, áëèçêèõ ê âåëè÷èíàì èõ ðÊ (ò.å. ïðè ðÍ 1,7—3,2 è 8,6-10,8). Ñòåðåîõèìèÿ àìèíîêèñëîò. Âàæíåéøèì ñâîéñòâîì àìèíîêèñëîò, îñâîáîæäàþùèõñÿ â ïðîöåññå ãèäðîëèçà ïðèðîäíûõ áåëêîâ â óñëîâèÿõ, èñêëþ÷àþùèõ ðàöåìèçàöèþ, ÿâëÿåòñÿ èõ îïòè÷åñêàÿ àêòèâíîñòü. Áóäó÷è ðàñòâîðåííûìè â âîäå (èëè â ÍÑ1), îíè ñïîñîáíû âðàùàòü ïëîñêîñòü ïîëÿðèçîâàííîãî ëó÷à (èñêëþ÷åíèå ñîñòàâëÿåò ãëèöèí). Ýòî ñâîéñòâî ñâÿçàíî ñ íàëè÷èåì â ìîëåêóëå âñåõ ïðèðîäíûõ àìèíîêèñëîò (çà èñëþ÷åíèåì ãëèöèíà) â à-ïîëîæåíèè àñèììåòðè÷åñêîãî àòîìà óãëåðîäà (ò. å. àòîìà óãëåðîäà, âñå ÷åòûðå âàëåíòíûå ñâÿçè êîòîðîãî çàíÿòû ðàçëè÷íûìè çàìåñòèòåëÿìè). Âåëè÷èíû óäåëüíîãî âðàùåíèÿ âïðàâî èëè âëåâî ÿâëÿþòñÿ êîëè÷åñòâåííîé õàðàêòåðèñòèêîé îïòè÷åñêîé àêòèâíîñòè, è äëÿ áîëüøèíñòâà àìèíîêèñëîò ñîñòàâëÿåò îò 10 äî 30°. Ïðèìåðíî ïîëîâèíà àìèíîêèñëîò áåëêîâ îêàçàëàñü ïðàâîâðàùàþùåé, èõ îáîçíà÷àþò çíàêîì «+» (Àëà, Èëå, Ãëó, Ëèç è äð.), à ÷óòü ìåíüøå ïîëîâèíû—ëåâîâðàùàþùåé (Ôåí, Òðï, Ëåé è äð.), èõ îáîçíà÷àþò çíàêîì «—». Âñå ýòè àìèíîêèñëîòû ïðèíàäëåæàò ê Ü-ðÿäó, à âåëè÷èíà è çíàê îïòè÷åñêîãî âðàùåíèÿ çàâèñÿò îò ïðèðîäû ðàäèêàëîâ àìèíîêèñëîò è çíà÷åíèÿ ðÍ ðàñòâîðà, â êîòîðîì èçìåðÿþò îïòè÷åñêîå âðàùåíèå. Ñòåðåîõèìèþ àìèíîêèñëîò ïðèíÿòî îöåíèâàòü íå ïî îïòè÷åñêîìó âðàùåíèþ, à èñõîäÿ èç àáñîëþòíîé êîíôèãóðàöèè âñåõ ÷åòûðåõ çàìåùàþùèõ ãðóïï, ðàñïîëîæåííûõ âîêðóã àñèììåòðè÷åñêîãî àòîìà óãëåðîäà â âåðøè-íàõ ìîäåëè òåòðàýäðà. Àáñîëþòíóþ êîíôèãóðàöèþ àìèíîêèñëîò ïðèíÿòî ñîîòíîñèòü ñòåðåîõèìè÷åñêè ñ ñîåäèíåíèåì, ïðîèçâîëüíî âçÿòûì äëÿ ñðàâíåíèÿ, à èìåííî ñ ãëèöåðèíîâûì àëüäåãèäîì, òàêæå ñîäåðæàùèì àñèììåòðè÷åñêèé àòîì óãëåðîäà. Íèæå ïðåäñòàâëåíû Ü- è Á-ñòåðåîèçîìåðû ãëèöåðèíîâîãî àëüäåãèäà. Ðÿäîì ïîêàçàíû ïðîñòðàíñòâåííûå êîíôèãóðàöèè Ü- è Á-àëàíèíà: îáðàçîì, ïðèðîäíûå àìèíîêèñëîòû èìåþò ïðîñòðàíñòâåííîå ðàñïîëîæåíèå, àíàëîãè÷íîå êîíôèãóðàöèè Ü-ãëèöåðèíîâîãî àëüäåãèäà. Ñëåäóåò åùå ðàç ïîä÷åðêíóòü, ÷òî ñèìâîëû Ü è Á îçíà÷àþò ïðèíàäëåæíîñòü äàííîé àìèíîêèñëîòû ïî ñâîåé ñòåðåîõèìè÷åñêîé êîíôèãóðàöèè ê Ü- èëè Á-ðÿäó, â òî âðåìÿ êàê çíàê «+» èëè «-» óêàçûâàåò íà íàïðàâëåíèå èçìåíåíèÿ ïëîñêîñòè ïîëÿðèçàöèè ñâåòîâîãî ëó÷à. Ñðåäè áåëêîâûõ àìèíîêèñëîò èìåþòñÿ äâå àìèíîêèñëîòû (òðåîíèí è èçîëåéöèí), êîòîðûå ñîäåðæàò ïî äâà àñèììåòðè÷åñêèõ àòîìà óãëåðîäà. Ñëåäîâàòåëüíî, åñëè íå â ïðèðîäå, òî, âî âñÿêîì ñëó÷àå, â ëàáîðàòîðèè âîçìîæíî ïîëó÷èòü ÷åòûðå ñòåðåîèçî- ìåðíûå ôîðìû ýòèõ àìèíîêèñëîò . Äëÿ òðåîíèíà èçâåñòíû âñå ÷åòûðå èçîìåðà. Åñëè óñëîâíî îáîçíà÷èòü ñèìâîëîì Ü âûäåëåííûé èç ïðèðîäíûõ áåëêîâ òðåîíèí, òî åãî çåðêàëüíîå îòîáðàæåíèå íàçûâàþò Á-òðåîíèíîì. Äâà äðóãèõ èçîìåðà, ïîëó÷èâøèõ íàèìåíîâàíèå äèàñòåðåîèçîìåðîâ, èëè àëëîôîðì, òàêæå ìîãóò èìåòü Ü- è Á-ôîðìû. Ñòðóêòóðíûå êîíôèãóðàöèè âñåõ ÷åòûðåõ ñòåðåîèçîìåðîâ òðåîíèíà ìîæíî ïðåäñòàâèòü ñëåäóþùèìè ôîðìóëàìè: ÑÎÎÍ 1 ÑÎÎÍ ñ-í 1 Í-Ñ-ÃØ2 ñ-í í-ñ-îí 1 1 ÑÍç 1 ÑÍç íî- 1_-òðåîíèí Ý-òðåîíèí 1-àëëîòðåîíèí Ý-àëëîòðåîíèí Êàê îòìå÷àëîñü, â áåëêîâîé ìîëåêóëå Á-àìèíîêèñëîòû íå îáíàðóæåíû , îäíàêî â æèâîé ïðèðîäå îíè øèðîêî ðàñïðîñòðàíåíû. Òàê, Á-èçîìåðû ãëóòàìèíîâîé êèñëîòû, àëàíèíà, âàëèíà, ôåíèëàëàíèíà, ëåéöèíà è ðÿäà äðóãèõ îòêðûòû â êëåòî÷íîé ñòåíêå áàêòåðèé; â ñîñòàâå íåêîòîðûõ àíòèáèîòèêîâ, â ÷àñòíîñòè àêòèíîìèöèíîâ, áàöèòðàöèíà, ãðàìè- öèäèíîâ À è 8, ñîäåðæàòñÿ àìèíîêèñëîòû Á-êîíôèãóðàöèè. Àìèíîêèñëîòíûé ñîñòàâ (êà÷åñòâåííûé è êîëè÷åñòâåííûé) ìíîãèõ òûñÿ÷ áåëêîâ, ïîëó÷åííûõ èç ðàçíûõ èñòî÷íèêîâ, âûÿñíåí (òàáë. 1.4). Ïðè àíàëèçå äàííûõ òàáë. 1.4 âèäåí ðÿä çàêîíîìåðíîñòåé. Íà äîëþ äèêàðáîíîâûõ àìèíîêèñëîò è èõ àìèäîâ â áîëüøèíñòâå áåëêîâ ïðèõîäèòñÿ äî 25-27% âñåõ àìèíîêèñëîò. Ýòè æå àìèíîêèñëîòû âìåñòå ñ ëåéöèíîì è ëèçèíîì ñîñòàâëÿþò îêîëî 50% âñåõ àìèíîêèñëîò. Â òî æå âðåìÿ íà äîëþ òàêèõ àìèíîêèñëîò, êàê öèñòåèí, ìåòèîíèí, òðèïòîôàí, ãèñòèäèí, ïðèõîäèòñÿ íå áîëåå 1,5-3,5%. Â ïðîòàìèíàõ è ãèñòîíàõ îòìå÷åíî âûñîêîå ñîäåðæàíèå îñíîâíûõ àìèíîêèñëîò àðãèíèíà è ëèçèíà, ñîîòâåòñòâåííî 26,4 è 85,2% (ñì. «Õèìèÿ ïðîñòûõ áåëêîâ»). Õèìè÷åñêèå ðåàêöèè äëÿ îòêðûòèÿ è îïðåäåëåíèÿ àìèíîêèñëîò â ãèäðîëèçàòàõ áåëêîâ. Â êóðñå îðãàíè÷åñêîé õèìèè ïîäðîáíî ðàññìîòðåíî ìíîæåñòâî õèìè÷åñêèõ ðåàêöèé, õàðàêòåðíûõ äëÿ à-àìèíî- è à-êàðáîêñèëüíûõ ãðóïï àìèíîêèñëîò (àöèëèðîâàíèå, àëêèëèðîâàíèå, íèòðîâàíèå, ýòåðèôèêàöèÿ Òàáëèöà 1.4. Àìèíîêèñëîòíûé ñîñòàâ íåêîòîðûõ ïðèðîäíûõ áåëêîâ, â ïðîöåíòàõ Áåëîê Àìèíîêèñëîòà Ñàëü- ìèí Ãèñòîí (ïå÷åíü òåëåí- êà) Êàçåèí Àëüáó-ìèí (ñû-âîðîòêà ÷åëîâåêà) ó-Ãëî- áóëèí (÷åëî âåêà) Ïåïñèí Èíñó ëèí Êîëëà- ãåí Àëàíèí 1,1 7,6 3,2 — — 4,5 9,5 Ãëèöèí 2,9 5,8 2,0 1,6 4,2 6,4 4,3 27,2 Âàëèí 3,1 5,5 7,2 7,7 9,7 7,1 7,7 3,4 Ëåéöèí 0 9,1 9,2 11,0 9,3 10,4 13,2 — Èçîëåéöèí 1,6 4,6 6,1 1,7 2,7 10,8 2,8 5,6 Ïðîëèí 5,8 3,4 10,6 5,1 8,1 5,0 2,5 15,1 Ôåíèëàëàíèí 0 3,5 5,0 7,8 4,6 6,4 8,8 2,5 Òèðîçèí 0 3,9 6,3 4,7 6,8 8,5 13,0 1,0 Òðèïòîôàí 0 - 1,2 0,2 2,9 2,4 0 0 Ñåðèí 9,1 4,1 6,3 3,3 11,4 12,2 5,2 3,4 Òðåîíèí 0 6,4 4,9 4,6 8,4 9,6 2,1 2,3 Öèñòåèí + öèñòèí 0 — 0,3 6,3 3,1 2,1 12,5 0 Ìåòèîíèí 0 0,9 2,8 1,3 1,1 1,7 — 0,8 Àðãèíèí 85,2 14,8 4,1 6,2 4,8 1,0 3,1 8,6 Ãèñòèäèí 0 2,3 3,1 3,5 2,5 0,9 4,9 0,7 Ëèçèí 0 11,7 8,2 12,3 8,1 0,9 2,5 4,5 Àñïàðàãèíîâàÿ êèñëîòà 0 5,5 7,1 9,0 8,8 16,0 6,8 6,3 Ãëóòàìèíîâàÿ êèñëîòà 0 10,3 22,4 17,0 11,8 1,1 11,9 18,6 11,3 Àìèäíûé àçîò 0 0,7 1,6 0,9 1,3 1,4 0,7 è äð.). Çäåñü áóäóò ðàññìîòðåíû îáùèå öâåòíûå ðåàêöèè äëÿ îáíàðóæåíèÿ èíäèâèäóàëüíûõ àìèíîêèñëîò è àìèíîêèñëîò, âõîäÿùèõ â ñîñòàâ áåëêîâ, îñíîâàííûå íà õèìè÷åñêîé ïðèðîäå ðàäèêàëîâ àìèíîêèñëîò (òàáë. 1.5). Äëÿ îòêðûòèÿ â áèîîáúåêòàõ è êîëè÷åñòâåííîãî îïðåäåëåíèÿ àìèíîêèñëîò óñïåøíî ïðèìåíÿåòñÿ ðåàêöèÿ èõ ñ íèíãèäðèíîì. Íà I ñòàäèè ðåàêöèè îáðàçóåòñÿ âîññòàíîâëåííûé íèíãèäðèí çà ñ÷åò îêèñëèòåëüíîãî äåçàìèíèðîâàíèÿ àìèíîêèñëîò (ïàðàëëåëüíî ïðîèñõîäèò äåêàðáîêñèëèðîâàíèå àìèíîêèñëîò): Íà II ñòàäèè îáðàçîâàâøèéñÿ àììèàê ðåàãèðóåò ñ ýêâèìîëÿðíûìè êîëè÷åñòâàìè îêèñëåííîãî è âîññòàíîâëåííîãî íèíãèäðèíà, îáðàçóÿ ñèíå-ôèîëåòîâûé ïðîäóêò, èíòåíñèâíîñòü îêðàñêè êîòîðîãî (ïðè 570 íì) ïðî-ïîðöèîíàëüíà êîëè÷åñòâó àìèíîêèñëîòû: Íà îñíîâå íèíãèäðèíîâîé ðåàêöèè áûëè ðàçðàáîòàíû ìåòîäû êîëè÷åñòâåííîãî îïðåäåëåíèÿ àìèíîêèñëîò, â ÷àñòíîñòè ìåòîä ðàñïðåäåëèòåëüíîé õðîìàòîãðàôèè íà áóìàãå, âïåðâûå âíåäðåííûé â 1944 ã. (À. Ìàðòèí è Ð. Ñèíäæ). Ýòà æå ðåàêöèÿ èñïîëüçóåòñÿ áëàãîäàðÿ ñâîåé âûñîêîé ÷óâñòâèòåëüíîñòè â àâòîìàòè÷åñêîì àíàëèçàòîðå àìèíîêèñëîò. Âïåðâûå òàêîé ïðèáîð ñêîíñòðóèðîâàëè Ä. Øïàêìàí, Ñ. Ìóð è Ó. Ñòåéí (ðèñ. 1.7). Ïîñëå ðàçäåëåíèÿ ñìåñè àìèíîêèñëîò â êîëîíêàõ, çàïîëíåííûõ ñïåöèàëüíûìè èîíîîáìåííûìè ñìîëàìè (ñóëüôîïîëèñòèðîëüíûé êàòèîíèò), òîê ýëþåíòà èç êîëîíêè ïîñòóïàåò â ñìåñèòåëü, òóäà æå ïîñòóïàåò ðàñòâîð íèíãèäðèíà; èíòåíñèâíîñòü îáðàçóþùåéñÿ îêðàñêè àâòîìàòè÷åñêè èçìåðÿåòñÿ íà ôîòîýëåêòðîêîëîðèìåòðå è ðåãèñòðèðóåòñÿ ñàìîïèñöåì. Ýòîò ìåòîä íàøåë øèðîêîå ïðèìåíåíèå â êëèíè÷åñêîé ïðàêòèêå ïðè èññëåäîâàíèè êðîâè, ìî÷è, ñïèííîìîçãîâîé æèäêîñòè. Ñ åãî ïîìîùüþ çà 2—3 ÷ ìîæíî ïîëó÷èòü ïîëíóþ êàðòèíó êà÷åñòâåííîãî ñîñòàâà àìèíîêèñëîò â áèîëîãè- Òàáëèöà 1.5. Ðåàêöèè, èñïîëüçóåìûå äëÿ èäåíòèôèêàöèè è ïîëóêîëè÷åñòâåííîãî îïðåäåëåíèÿ àìèíîêèñëîò è áåëêîâ Ðåàêöèÿ Ðåàêòèâû Îïðåäåëÿåìàÿ àìèíîêèñëîòà Îêðàñêà Ìèëëîíà ÍäÌ03 â àçîòíîé êèñëîòå â ïðè-ñóòñòâèè àçîòèñòîé êèñëîòû Òèðîçèí Êðàñíàÿ Êñàíòîïðîòåè- íîâàÿ Êèïÿùàÿ êîíöåíòðèðîâàííàÿ àçîòíàÿ êèñëîòà Ôåíèëàëàíèí, òèðîçèí Æåëòàÿ Ãîïêèíñà- Êîóëà Ãëèîêñèëîâàÿ êèñëîòà â êîí-öåíòðèðîâàííîé ñåðíîé êèñëîòå Òðèïòîôàí Ñèíå-ôèîëå- òîâàÿ Ýðëèõà ï-Äèìåòèëàìèíîáåíçàëüäåãèä â êîíöåíòðèðîâàííîé õëîðèñ-òîâîäîðîäíîé êèñëîòå Òðèïòîôàí Ñèíÿÿ Ñàêàãó÷è à-Íàôòîë è ãèïîõëîðèò íàòðèÿ Àðãèíèí Êðàñíàÿ Íèòðîïðóñ- ñèäíàÿ Íèòðîïðóññèä íàòðèÿ â ðàçáàâ-ëåííîì ðàñòâîðå àììèàêà Öèñòåèí Êðàñíàÿ Ñàëëèâåíà 1,2-Íàôòîõèíîí-4-ñóëüôîíàò íàòðèÿ è áèñóëüôèò íàòðèÿ Öèñòåèí » Ïàóëè Äèàçîòèðîâàííàÿ ñóëüôàíèëî- âàÿ êèñëîòà â ùåëî÷íîì ðàñòâîðå Ãèñòèäèí, òèðîçèí » Ôîëèíà- ×îêàëòåó Ôîñôîìîëèáäåíîâî-âîëüôðà- ìîâàÿ êèñëîòà Òèðîçèí Ñèíÿÿ Ðèñ. 1.7. Ðàáîòà àâòîìàòè÷åñêîãî àíàëèçàòîðà àìèíîêèñëîò (ïðèíöèïèàëüíàÿ ñõåìà ïî Øïàêìàíó, Ìóðó è Ñòåéíó). 1 - ñìåñèòåëü; 2 - ôîòîýëåêòðîêîëîðèìåòð; 3 - ñàìîïèñåö. ÷åñêèõ æèäêîñòÿõ è âûÿâèòü íàëè÷èå â íèõ íåîáû÷íûõ àçîòñîäåðæàùèõ âåùåñòâ, ÷òî èìååò âàæíîå äèàãíîñòè÷åñêîå è ïðîãíîñòè÷åñêîå çíà÷åíèå. Àâòîìàòè÷åñêèå àíàëèçàòîðû àìèíîêèñëîò âñå âðåìÿ ñîâåðøåíñòâóþòñÿ, ïîâûøàþòñÿ ÷óâñòâèòåëüíîñòü ìåòîäîâ è ñêîðîñòü ïðîâåäåíèÿ àíàëèçà. Òàê, â ñîâðåìåííûõ ïðèáîðàõ âûñîêîýôôåêòèâíîé æèäêîñòíîé õðîìàòîãðàôèè (ÂÝÆÕ) óäàåòñÿ ïðîâîäèòü àíàëèç ãèäðîëèçàòà áåëêà çà 45 ìèí, îïðåäåëÿÿ ïðè ýòîì êîíöåíòðàöèþ àìèíîêèñëîò â ïèêîìîëÿõ (ðèñ. 1.8). Ñìåñü àìèíîêèñëîò ìîæåò áûòü óñïåøíî ðàçäåëåíà òàêæå ìåòîäîì ýëåêòðîôîðåçà íà áóìàãå. Ïðè ðÍ 6,0 âîçìîæíî õîðîøåå ðàçäåëåíèå êèñëûõ è îñíîâíûõ àìèíîêèñëîò ñ íåéòðàëüíûìè. Â ýòîì ñëó÷àå îòðèöàòåëüíî çàðÿæåííûå (êèñëûå) àìèíîêèñëîòû áóäóò äâèãàòüñÿ ê àíîäó, à ïîëîæèòåëüíî çàðÿæåííûå—ê êàòîäó. Íåéòðàëüíûå àìèíîêèñëîòû îñòàþòñÿ íà ëèíèè ñòàðòà. Äëÿ èõ ðàçäåëåíèÿ ýëåêòðîôîðåç îáû÷íî ïðîâîäÿò ïðè ðÍ 1,8—2,0, êîãäà âñå îíè ìèãðèðóþò ê àíîäó ñ íåçíà÷èòåëüíûì, íî óëîâèìûì ðàçëè÷èåì â ïîäâèæíîñòè. Ïîñëå ýëåêòðîôîðåçà ìåñòîïîëîæåíèå àìèíîêèñëîò íà ýëåêòîôîðåãðàììå âûÿâëÿþò ñ ïîìîùüþ õèìè÷åñêèõ ðåàêöèé, à ïîñëå ýëþöèè îêðàøåííûõ ïðîäóêòîâ îïðåäåëÿþò èõ êîëè÷åñòâåííî. Ðèñ. 1.8. ÂÝÆÕ àìèíîêèñëîò ïî Öåõó è Âîëüòåðó. Ðàçäåëåíèå íà êîëîíêå (3 õ 250 ìì), íàïîëíåííîé èîíîîáìåííîé ñìîëîé - ïîëè- ñòèðîëäèâèíèëáåíçîëîì. Êîíöåíò-ðàöèÿ àìèíîêèñëîò 500 ïìîëü/ë, ðåàêòèâ äëÿ äåòåêòèðîâàíèÿ - ôëþîðåñêàìèí, îáðàçóþùèé ñ àìèíîãðóïïîé ñèëüíî ôëþîðåñöèðóþùåå ñîåäèíåíèå. 1 - Àñï; 2 - Òðå; 3 - Ñåð; 4 - Ãëó; 5 - Ãëè; 6 - Àëà; 7 - Öèñ; 8 - Âàë; 9 - Ìåò; 10 - Èëå; 11 - Ëåé; 12 - Òèð; 13 - Ôåí; 14 - Ëèç; 15 - Ãèñ; 16 - Àðã. ÔÈÇÈÊÎ-ÕÈÌÈ×ÅÑÊÈÅ ÑÂÎÉÑÒÂÀ ÁÅËÊÎÂ Íàèáîëåå õàðàêòåðíûìè ôèçèêî-õèìè÷åñêèìè ñâîéñòâàìè áåëêîâ ÿâëÿþòñÿ âûñîêàÿ âÿçêîñòü ðàñòâîðîâ, íåçíà÷èòåëüíàÿ äèôôóçèÿ, ñïîñîáíîñòü ê íàáóõàíèþ â áîëüøèõ ïðåäåëàõ, îïòè÷åñêàÿ àêòèâíîñòü, ïîäâèæíîñòü â ýëåêòðè÷åñêîì ïîëå, íèçêîå îñìîòè÷åñêîå äàâëåíèå è âûñîêîå îíêîòè÷åñêîå äàâëåíèå, ñïîñîáíîñòü ê ïîãëîùåíèþ ÓÔ-ëó÷åé ïðè 280 íì (ýòî ñâîéñòâî, îáóñëîâëåííîå íàëè÷èåì â áåëêàõ àðîìàòè÷åñêèõ àìèíîêèñëîò, èñïîëüçóåòñÿ äëÿ êîëè÷åñòâåííîãî îïðåäåëåíèÿ áåëêîâ). Áåëêè, êàê è àìèíîêèñëîòû, àìôîòåðíû áëàãîäàðÿ íàëè÷èþ ñâîáîäíûõ ÌÈ2- è ÑÎÎÍ-ãðóïï. Äëÿ íèõ õàðàêòåðíû âñå ñâîéñòâà êèñëîò è îñíîâàíèé. Â çàâèñèìîñòè îò ðåàêöèè ñðåäû è ñîîòíîøåíèÿ êèñëûõ è îñíîâíûõ àìèíîêèñëîò áåëêè â ðàñòâîðå íåñóò èëè îòðèöàòåëüíûé, èëè ïîëîæèòåëüíûé çàðÿä, ïåðåìåùàÿñü ê àíîäó èëè êàòîäó. Ýòî ñâîéñòâî èñïîëüçóåòñÿ ïðè î÷èñòêå áåëêîâ ìåòîäîì ýëåêòðîôîðåçà. Áåëêè îáëàäàþò ÿâíî âûðàæåííûìè ãèäðîôèëüíûìè ñâîéñòâàìè. Ðàñòâîðû áåëêîâ èìåþò î÷åíü íèçêîå îñìîòè÷åñêîå äàâëåíèå, âûñîêóþ âÿçêîñòü è íåçíà÷èòåëüíóþ ñïîñîáíîñòü ê äèôôóçèè. Áåëêè ñïîñîáíû ê íàáóõàíèþ â î÷åíü áîëüøèõ ïðåäåëàõ. Ñ êîëëîèäíûì ñîñòîÿíèåì áåëêîâ ñâÿçàí ðÿä õàðàêòåðíûõ ñâîéñòâ, â ÷àñòíîñòè ÿâëåíèå ñâåòîðàññåÿíèÿ, ëåæàùåå â îñíîâå êîëè÷åñòâåííîãî îïðåäåëåíèÿ áåëêîâ ìåòîäîì íåôåëîìåòðèè. Ýòîò ýôôåêò èñïîëüçóåòñÿ, êðîìå òîãî, â ñîâðåìåííûõ ìåòîäàõ ìèêðîñêîïèè áèîëîãè÷åñêèõ îáúåêòîâ. Ìîëåêóëû áåëêà íå ñïîñîáíû ïðîíèêàòü ÷åðåç ïîëóïðîíèöàåìûå èñêóññòâåííûå ìåìáðàíû (öåëëîôàí, ïåðãàìåíò, êîëëîäèé), à òàêæå áèîìåìáðàíû ðàñòèòåëüíûõ è æèâîòíûõ òêàíåé, õîòÿ ïðè îðãàíè÷åñêèõ ïîðàæåíèÿõ, íàïðèìåð, ïî÷åê êàïñóëà ïî÷å÷íîãî êëóáî÷êà (Øóìëÿíñêîãî- Áîóìåíà) ñòàíîâèòñÿ ïðîíèöàåìîé äëÿ àëüáóìèíîâ ñûâîðîòêè êðîâè è ïîñëåäíèå ïîÿâëÿþòñÿ â ìî÷å. Ìîëåêóëÿðíàÿ ìàññà áåëêîâ Áåëêè îòíîñÿòñÿ ê âûñîêîìîëåêóëÿðíûì ñîåäèíåíèÿì, â ñîñòàâ êîòîðûõ âõîäÿò ñîòíè è äàæå òûñÿ÷è àìèíîêèñëîòíûõ îñòàòêîâ, îáúåäèíåííûõ â ìàêðîìîëåêóëÿðíóþ ñòðóêòóðó. Ìîëåêóëÿðíàÿ ìàññà áåëêîâ êîëåáëåòñÿ îò 6000 (íèæíèé ïðåäåë) äî 1000000 è âûøå â çàâèñèìîñòè îò êîëè÷åñòâà îòäåëüíûõ ïîëèïåïòèäíûõ öåïåé â ñîñòàâå åäèíîé ìîëåêóëÿðíîé ñòðóêòóðû áåëêà. Òàêèå ïîëèïåïòèäíûå öåïè ïîëó÷èëè íàçâàíèå ñóáúåäèíèö. Èõ ìîë. ìàññà âàðüèðóåò â øèðîêèõ ïðåäåëàõ—îò 6000 äî 100000 è áîëåå. Àìèíîêèñëîòíûé ñîñòàâ è ïîñëåäîâàòåëüíîñòü àìèíîêèñëîò âûÿñíåíû äëÿ ìíîãèõ òûñÿ÷ áåëêîâ. Â ñâÿçè ñ ýòèì ñòàëî âîçìîæíûì âû÷èñëåíèå èõ ìîëåêóëÿðíîé ìàññû õèìè÷åñêèì ïóòåì ñ âûñîêîé òî÷íîñòüþ. Îäíàêî äëÿ îãðîìíîãî êîëè÷åñòâà âñòðå÷àþùèõñÿ â ïðèðîäå áåëêîâ õèìè÷åñêîå ñòðîåíèå íå âûÿñíåíî, ïîýòîìó îñíîâíûìè ìåòîäàìè îïðåäåëåíèÿ ìîëåêóëÿðíîé ìàññû âñå åùå îñòàþòñÿ ôèçèêî-õèìè÷åñêèå ìåòîäû (ãðàâèìåòðè÷åñêèå, îñìîìåòðè÷åñêèå, âèñêîçèìåòðè÷åñêèå, ýëåêòðîôîðåòè÷åñêèå, îïòè÷åñêèå è äð.). Íà ïðàêòèêå íàèáîëåå ÷àñòî èñïîëüçóþòñÿ ìåòîäû ñåäèìåíòàöèîííîãî àíàëèçà, ãåëü-õðîìàòîãðàôèÿ è ãåëü-ýëåêòðîôîðåç. Îïðåäåëåíèå ìîëåêóëÿðíîé ìàññû áåëêîâ ìåòîäàìè ñåäèìåíòàöèîííîãî àíàëèçà ïðîâîäÿò â óëüòðàöåíòðèôóãàõ , â êîòîðûõ óäàåòñÿ ñîçäàòü öåíòðîáåæíûå óñêîðåíèÿ (§), ïðåâûøàþùèå â 200000 è áîëåå ðàç óñêîðåíèå çåìíîãî ïðèòÿæåíèÿ. Îáû÷íî âû÷èñëÿþò ìîëåêóëÿðíóþ ìàññó ïî ñêîðîñòè ñåäèìåíòàöèè ìîëåêóë áåëêà èëè ñåäèìåíòàöèîííîìó ðàâíîâåñèþ. Ïî ìåðå ïåðåìåùåíèÿ ìîëåêóë îò öåíòðà ê ïåðèôåðèè îáðàçóåòñÿ ðåçêàÿ ãðàíèöà ðàñòâîðèòåëü- áåëîê (ðåãèñòðèðóåòñÿ àâòîìàòè÷åñêè). Îïòè÷åñêèå ñâîéñòâà ðàñòâîðèòåëÿ è áåëêà èñïîëüçóþòñÿ ïðè îïðåäåëåíèè ñêîðîñòè ñåäèìåíòàöèè; ïîñëåäíþþ âûðàæàþò ÷åðåç êîíñòàíòó ñåäèìåíòàöèè 8, êîòîðàÿ çàâèñèò êàê îò ìàññû, òàê è îò ôîðìû áåëêîâîé ÷àñòèöû: V ãäå V—ñêîðîñòü ïåðåìåùåíèÿ ãðàíèöû ðàñòâîðèòåëü-áåëîê, ñì/ñ; þ — óãëîâàÿ ñêîðîñòü ðîòîðà, ðàä/ñ; ã — ðàññòîÿíèå îò öåíòðà ðîòîðà äî ñåðåäèíû ÿ÷åéêè ñ ðàñòâîðîì áåëêà, ñì. Êîíñòàíòà ñåäèìåíòàöèè èìååò ðàçìåðíîñòü âðåìåíè (åå âûðàæàþò â ñåêóíäàõ). Âåëè÷èíà êîíñòàíòû ñåäèìåíòàöèè, ðàâíàÿ 1 • 10-13 ñ, óñëîâíî ïðèíÿòà çà åäèíèöó è íàçâàíà ñâåäáåðãîì (8). Çíà÷åíèÿ êîíñòàíò ñåäèìåíòàöèè áîëüøèíñòâà áåëêîâ ëåæàò â ïðåäåëàõ 1—50 8, õîòÿ â ðÿäå ñëó÷àåâ ýòè çíà÷åíèÿ ïðåâûøàþò 100 8. Äëÿ âû÷èñëåíèÿ ìîëåêóëÿðíîé ìàññû (Ì), ïîìèìî êîíñòàíòû ñåäèìåíòàöèè, íåîáõîäèìû äîïîëíèòåëüíûå ñâåäåíèÿ î ïëîòíîñòè ðàñòâîðèòåëÿ è áåëêà è äðóãèå ñîãëàñíî óðàâíåíèþ Ñâåäáåðãà: ãäå Ê — ãàçîâàÿ ïîñòîÿííàÿ, ýðã/(ìîëü • ãðàä); Ò — àáñîëþòíàÿ òåìïåðàòóðà (ïî øêàëå Êåëüâèíà); ÿ—êîíñòàíòà ñåäèìåíòàöèè; ð — ïëîòíîñòü ðàñòâîðèòåëÿ; V — ïàðöèàëüíûé óäåëüíûé îáúåì ìîëåêóëû áåëêà; Á - êîýôôèöèåíò äèôôóçèè. Îïðåäåëåíèå ìîëåêóëÿðíîé ìàññû áåëêîâ ìåòîäîì óëüòðàöåíòðèôóãèðîâàíèÿ òðåáóåò ìíîãî âðåìåíè è ñëîæíîé è äîðîãîñòîÿùåé àïïàðàòóðû. Ïîýòîìó â ïîñëåäíèå ãîäû ðàçðàáîòàíû äâà áîëåå ïðîñòûõ ìåòîäà (ãåëü- õðîìàòîãðàôèÿ è ýëåêòðîôîðåç). Ïðè èñïîëüçîâàíèè ãåëü-õðîìàòîãðàôèè â ïåðâóþ î÷åðåäü òðåáóåòñÿ îòêàëèáðîâàòü êîëîíêó. Äëÿ ýòîãî ÷åðåç êîëîíêó ñ ñåôàäåêñîì ïðîïóñêàþò íåñêîëüêî áåëêîâ ñ èçâåñòíûìè ìîëåêóëÿðíûìè ìàññàìè è ñòðîÿò ãðàôèê, îòêëàäûâàÿ çíà÷åíèÿ ëîãàðèôìîâ ìîëåêóëÿðíîé ìàññû ïðîòèâ èõ ýëþöèîííûõ îáúåìîâ, êîòîðûå íàõîäÿò, êàê ïîêàçàíî íà ðèñ. 1.9. Èçâåñòíî, ÷òî ìåæäó ëîãàðèôìîì ìîëåêóëÿðíîé ìàññû áåëêà, èìåþùåãî ñôåðè÷åñêóþ ôîðìó, è ýëþöèîííûì îáúåìîì ñóùåñòâóåò ïðÿìàÿ çàâèñèìîñòü. Ïîýòîìó ëåãêî îïðåäåëèòü ìîëåêóëÿðíóþ ìàññó èññëåäóåìîãî áåëêà, çíàÿ åãî îáúåì ýëþöèè. Âòîðîé ðàçíîâèäíîñòüþ ýòîãî ìåòîäà ÿâëÿåòñÿ òîíêîñëîéíàÿ ãåëü-õðîìàòîãðàôèÿ. Äëèíà ïðîáåãà áåëêà (â ìèëëèìåòðàõ) Ðèñ. 1.9. Èçìåðåíèå îáúåìà ýëþöèè (ó^)- Ðèñ. 1.10. Çàâèñèìîñòü ìåæäó äëèíîé ïðîáåãà áåëêîâûõ ÷àñòèö ïðè ãåëü-õðîìàòîãðàôèè â òîíêîì ñëîå ñåôàäåêñà Ã-150 (ñâåðõòîíêîãî) è èõ ìîëåêóëÿðíûìè ìàññàìè (â ïîëóëîãàðèôìè÷åñêîé ñèñòåìå êîîðäèíàò). 1 - ðèáîíóêëåàçà; 2 - õèìîòðèïñèíîãåí; 3 - ÿè÷íûé àëüáóìèí; 4 - ñûâîðîòî÷íûé àëüáóìèí; 5 - ó-ãëîáóëèí; Õ - áåëîê ñ íåèçâåñòíîé ìîëåêóëÿðíîé ìàññîé. ÷åðåç òîíêèé ñëîé ñåôàäåêñà íàõîäèòñÿ â ëîãàðèôìè÷åñêîé çàâèñèìîñòè îò ìîëåêóëÿðíîé ìàññû áåëêà (ðèñ. 1.10). Ãåëü-õðîìàòîãðàôèÿ, êðîìå ïðîñòîòû è áûñòðîòû, èìååò äîïîëíèòåëüíîå ïðåèìóùåñòâî: íå òðåáóåòñÿ âûäåëÿòü áåëîê â ÷èñòîì âèäå, òàê êàê ïðèìåñè äðóãèõ áåëêîâ íå ìåøàþò îïðåäåëåíèþ, ïîñêîëüêó êàæäûé èç íèõ ïðîõîäèò ÷åðåç êîëîíêó ñî ñâîéñòâåííîé åìó ñêîðîñòüþ, îïðåäåëÿåìîé ìîëåêóëÿðíîé ìàññîé. Ýòî îáñòîÿòåëüñòâî øèðîêî èñïîëüçóåòñÿ â ýíçèìî- ëîãèè, êîãäà îêàçûâàåòñÿ âîçìîæíûì îïðåäåëåíèå ìîëåêóëÿðíîé ìàññû äàæå î÷åíü íåáîëüøîãî êîëè÷åñòâà ôåðìåíòà â ïðèñóòñòâèè äðóãèõ áåëêîâ, íå îáëàäàþùèõ àíàëîãè÷íîé êàòàëèòè÷åñêîé àêòèâíîñòüþ. Ïðè èñïîëüçîâàíèè äèñê-ýëåêòðîôîðåçà â ïîëèàêðèëàìèäíîì ãåëå äëÿ îïðåäåëåíèÿ ìîëåêóëÿðíîé ìàññû áåëêîâ òàêæå ñòðîÿò ãðàôèê çàâèñèìîñòè ìåæäó ëîãàðèôìîì ìîëåêóëÿðíîé ìàññû êàëèáðîâî÷íûõ áåëêîâ è ïîäâèæíîñòüþ áåëêîâûõ ÷àñòèö â ïîëèàêðèëàìèäíîì ãåëå, à çàòåì, îïðåäåëèâ ïîäâèæíîñòü èññëåäóåìîãî áåëêà, ïî ãðàôèêó íàõîäÿò åãî ìàññó (ðèñ. 1.11). Ýëåêòðîôîðåç ïðîâîäÿò â ïðèñóòñòâèè äåòåðãåíòà äîäåöèëñóëüôàòà íàòðèÿ, òàê êàê òîëüêî â ýòîì ñëó÷àå íàáëþäàåòñÿ ïðÿìàÿ ïðîïîðöèîíàëüíàÿ çàâèñèìîñòü ìåæäó ìîëåêóëÿðíîé ìàññîé è ïîäâèæíîñòüþ áåëêîâ. Áåëêè ñ ÷åòâåðòè÷íîé ñòðóêòóðîé ïðè ýòèõ óñëîâèÿõ ðàñïàäàþòñÿ íà ñóáúåäèíèöû, ïîýòîìó ìåòîä íàõîäèò øèðîêîå ïðèìåíåíèå äëÿ îïðåäåëåíèÿ ìîëåêóëÿðíîé ìàññû ñóáúåäèíèö áåëêà. 0,2 0,4 0,6 0,8 Îòíîñèòåëüíàÿ ïîäâèæíîñòü Ðèñ. 1.11. Çàâèñèìîñòü ìåæäó ìîëåêóëÿðíîé ìàññîé è îòíîñèòåëüíîé ïîäâèæíîñòüþ áåëêà ïðè äèñê-ýëåêòðîôîðåçå â ïîëèàêðèëàìèäíîì ãåëå â ïðèñóòñòâèè äîäåöèëñóëüôàòà íàòðèÿ (â ïîëóëîãàðèôìè÷åñêîé ñèñòåìå êîîðäèíàò). 1 - ñûâîðîòî÷íûé àëüáóìèí; 2 - ÿè÷íûé àëüáóìèí; 3 - ïåïñèí; 4 - õèìîòðèïñèíîãåí; 5 - ìèî- ãëîáèí; 6 - öèòîõðîì ñ; Õ - áåëîê ñ íåèçâåñòíîé ìîëåêóëÿðíîé ìàññîé. Íåäàâíî ïðåäëîæåí íîâûé ìàññ-ñïåêòðîìåòðè÷åñêèé ìåòîä (òàê íàçûâàåìûé ëàçåðíûé äåñîðáöèîííî-èîíèçàöèîííûé ìåòîä), ïîçâîëÿþùèé îïðåäåëÿòü ìîëåêóëÿðíóþ ìàññó íåáîëüøèõ ïåïòèäîâ (âàçîïðåññèí, èíñóëèí) è êðóïíûõ áèîïîëèìåðíûõ ìîëåêóë è, êðîìå òîãî, ñòðóêòóðó áèîìîëåêóë. Ôîðìà áåëêîâûõ ìîëåêóë Î âåëè÷èíå è ôîðìå áåëêîâûõ ìîëåêóë ðàíüøå ñóäèëè ïî äàííûì óëüòðàöåíòðèôóãèðîâàíèÿ, äâîéíîãî ëó÷åïðåëîìëåíèÿ è äèôôóçèè. Ýòè äàííûå óêàçûâàëè íà ñóùåñòâîâàíèå â ïðèðîäå ãëîáóëÿðíûõ (øàðîîáðàçíûõ) è ôèáðèëëÿðíûõ (íèòåâèäíûõ) áåëêîâ. Â íàñòîÿùåå âðåìÿ îáùèå ïðåäñòàâëåíèÿ î ôîðìå áåëêîâûõ ìîëåêóë â îñíîâíîì ïîäòâåðäèëèñü, îäíàêî òîëüêî ñîâðåìåííûå ìåòîäû èññëåäîâàíèÿ ïîçâîëèëè óñòàíîâèòü äåòàëè ïðîñò-ðàíñòâåííîé êîíôèãóðàöèè (òðåõìåðíîé ñòðóêòóðû) áåëêîâûõ ìîëåêóë. Áëàãîäàðÿ ïðèìåíåíèþ ñêàíèðóþùåé ìèêðîñêîïèè è ðåíòãåíîñòðóêòóðíîãî àíàëèçà (âûñîêîå ðàçðåøåíèå, ïîðÿäêà 0,2—0,3 íì) óäàëîñü â äåòàëÿõ ðàñøèôðîâàòü íå òîëüêî ïîëíóþ ïðîñòðàíñòâåííóþ ñòðóêòóðó, ôîðìó, íî è ñòåïåíü àñèììåòðèè áåëêîâûõ ìîëåêóë âî âñåõ òðåõ èçìåðåíèÿõ. Îêàçàëîñü, ÷òî äàæå ãëîáóëÿðíûå áåëêè êðîâè (ãåìîãëîáèí, àëüáóìèíû è ãëîáóëèíû) ÿâëÿþòñÿ àñèììåòðè÷íûìè â óêàçàííûõ èçìåðåíèÿõ. Ñëåäóåò îòìåòèòü, ÷òî íå òîëüêî ôèçèêî-õèìè÷åñêèå, íî è áèîëîãè÷åñêèå ñâîéñòâà áåëêîâ (â ñâîáîäíîì èëè â ñâÿçàííîì äðóã ñ äðóãîì èëè ñ äðóãèìè áèîïîëèìåðàìè ñîñòîÿíèè) îïðåäåëÿþòñÿ èõ ïðîñòðàíñòâåííîé ñòðóêòóðîé. Äåíàòóðàöèÿ áåëêîâ Ïðèðîäíûå áåëêîâûå òåëà íàäåëåíû îïðåäåëåííîé, ñòðîãî çàäàííîé ïðîñòðàíñòâåííîé êîíôèãóðàöèåé è îáëàäàþò ðÿäîì õàðàêòåðíûõ ôèçèêî-õèìè÷åñêèõ è áèîëîãè÷åñêèõ ñâîéñòâ ïðè ôèçèîëîãè÷åñêèõ çíà÷åíèÿõ òåìïåðàòóðû è ðÍ ñðåäû. Ïîä âëèÿíèåì ðàçëè÷íûõ ôèçè÷åñêèõ è õèìè÷åñêèõ ôàêòîðîâ áåëêè ïîäâåðãàþòñÿ ñâåðòûâàíèþ è âûïàäàþò â îñàäîê, òåðÿÿ íàòèâíûå ñâîéñòâà. Òàêèì îáðàçîì, ïîä äåíàòóðàöèåé ñëåäóåò ïîíèìàòü íàðóøåíèå îáùåãî ïëàíà óíèêàëüíîé ñòðóêòóðû íàòèâíîé ìîëåêóëû áåëêà, ïðåèìóùåñòâåííî åå òðåòè÷íîé ñòðóêòóðû, ïðèâîäÿùåå ê ïîòåðå õàðàêòåðíûõ äëÿ íåå ñâîéñòâ (ðàñòâîðèìîñòü, ýëåêòðîôîðåòè÷åñêàÿ ïîäâèæíîñòü, áèîëîãè÷åñêàÿ àêòèâíîñòü è ò.ä.). Áîëüøèíñòâî áåëêîâ äåíàòóðèðóåò ïðè íàãðåâàíèè èõ ðàñòâîðîâ âûøå 50—60°Ñ. Âíåøíèå ïðîÿâëåíèÿ äåíàòóðàöèè ñâîäÿòñÿ ê ïîòåðå ðàñòâîðèìîñòè, îñîáåííî â èçîýëåêòðè÷åñêîé òî÷êå, ïîâûøåíèþ âÿçêîñòè áåëêîâûõ ðàñòâîðîâ, óâåëè÷åíèþ êîëè÷åñòâà ñâîáîäíûõ ôóíêöèîíàëüíûõ 8Í-ãðóïï è èçìåíåíèþ õàðàêòåðà ðàññåèâàíèÿ ðåíòãåíîâñêèõ ëó÷åé. Íàèáîëåå õàðàêòåðíûì ïðèçíàêîì äåíàòóðàöèè ÿâëÿåòñÿ ðåçêîå ñíèæåíèå èëè ïîëíàÿ ïîòåðÿ áåëêîì åãî áèîëîãè÷åñêîé àêòèâíîñòè (êàòàëèòè÷åñêîé, àíòèãåííîé èëè ãîðìîíàëüíîé). Ïðè äåíàòóðàöèè áåëêà, âûçâàííîé 8Ì ìî÷åâèíîé èëè äðóãèì àãåíòîì, ðàçðóøàþòñÿ â îñíîâíîì íåêîâàëåíòíûå ñâÿçè (â ÷àñòíîñòè, ãèäðîôîáíûå âçàèìîäåéñòâèÿ è âîäîðîäíûå ñâÿçè). Äèñóëüôèäíûå ñâÿçè â ïðèñóòñòâèè âîññòàíàâëèâàþùåãî àãåíòà ìåðêàïòîýòàíîëà ðàçðûâàþòñÿ, â òî âðåìÿ êàê ïåïòèäíûå ñâÿçè ñàìîãî îñòîâà ïîëèïåïòèäíîé öåïè íå çàòðàãèâàþòñÿ. Â ýòèõ óñëîâèÿõ ðàçâåðòûâàþòñÿ ãëîáóëû íàòèâíûõ áåëêîâûõ ìîëåêóë è îáðàçóþòñÿ ñëó÷àéíûå è áåñïîðÿäî÷íûå ñòðóêòóðû (ðèñ. 1.12). Ïðè íåïðîäîëæèòåëüíîì äåéñòâèè è áûñòðîì óäàëåíèè äåíàòóðèðóþùèõ àãåíòîâ âîçìîæíà ðåíàòóðàöèÿ áåëêà ñ ïîëíûì âîññòàíîâëåíèåì à - èñõîäíîå ñîñòîÿíèå; á - íà÷èíàþùååñÿ îáðàòèìîå íàðóøåíèå ìîëåêóëÿðíîé ñòðóêòóðû; â - íåîáðàòèìîå ðàçâåðòûâàíèå ïîëè- ïåïòèäíîé öåïè. Ðèñ. 1.13. Äåíàòóðàöèÿ è ðå- íàòóðàöèÿ ðèáîíóêëåàçû (ïî Àíôèíñåíó). à - ðàçâåðòûâàíèå (ìî÷åâèíà + ìåðêàïòîýòàíîë); á - ïîâòîðíîå ñâåðòûâàíèå. èñõîäíîé òðåõìåðíîé ñòðóêòóðû è íàòèâíûõ ñâîéñòâ åãî ìîëåêóëû (ðèñ. 1.13), âêëþ÷àÿ áèîëîãè÷åñêóþ àêòèâíîñòü. Òàêèì îáðàçîì, ïðè äåíàòóðàöèè áåëêîâàÿ ìîëåêóëà ïîëíîñòüþ òåðÿåò áèîëîãè÷åñêèå ñâîéñòâà, äåìîíñòðèðóÿ òåì ñàìûì òåñíóþ ñâÿçü ìåæäó ñòðóêòóðîé è ôóíêöèåé. Äëÿ ïðàêòè÷åñêèõ öåëåé èíîãäà èñïîëüçóþò ïðîöåññ äåíàòóðàöèè â «ìÿãêèõ» óñëîâèÿõ, íàïðèìåð ïðè ïîëó÷åíèè ôåðìåíòîâ èëè äðóãèõ áèîëîãè÷åñêè àêòèâíûõ áåëêîâûõ ïðåïàðàòîâ â óñëîâèÿõ íèçêèõ òåìïåðàòóð â ïðèñóòñòâèè ñîëåé è ïðè ñîîòâåòñòâóþùåì çíà÷åíèè ðÍ . Ïðè ëèîôèëèçàöèè áåëêîâ (âûñóøèâàíèå â âàêóóìå ïóòåì âîçãîíêè âëàãè èç çàìîðîæåííîãî ñîñòîÿíèÿ) äëÿ ïðåäîòâðàùåíèÿ äåíàòóðàöèè ÷àñòî ïîëüçóþòñÿ õèìè÷åñêèìè âåùåñòâàìè (ïðîñòûå ñàõàðà, ãëèöåðèí, îðãàíè÷åñêèå àíèîíû). Èçîýëåêòðè÷åñêàÿ è èçîèîííàÿ òî÷êè áåëêîâ Â èçîýëåêòðè÷åñêîé òî÷êå ñóììàðíûé çàðÿä áåëêîâ, îáëàäàþùèõ àìôîòåð- íûìè ñâîéñòâàìè, ðàâåí íóëþ è áåëêè íå ïåðåìåùàþòñÿ â ýëåêòðè÷åñêîì ïîëå. Çíàÿ àìèíîêèñëîòíûé ñîñòàâ áåëêà, ìîæíî ïðèáëèæåííî îïðåäåëèòü èçîýëåêòðè÷åñêóþ òî÷êó (ð1); ð1 ÿâëÿåòñÿ õàðàêòåðíîé êîíñòàíòîé áåëêîâ. Èçîýëåêòðè÷åñêàÿ òî÷êà áîëüøèíñòâà áåëêîâ æèâîòíûõ òêàíåé ëåæèò â ïðåäåëàõ îò 5,5 äî 7,0, ÷òî ñâèäåòåëüñòâóåò î ÷àñòè÷íîì ïðåîáëàäàíèè êèñëûõ àìèíîêèñëîò. Îäíàêî â ïðèðîäå èìåþòñÿ áåëêè, ó êîòîðûõ çíà÷åíèÿ èçîýëåêòðè÷åñêèõ òî÷åê ëåæàò â êðàéíèõ çíà÷åíèÿõ ðÍ ñðåäû. Â ÷àñòíîñòè, âåëè÷èíà ð1 ïåïñèíà (ôåðìåíò æåëóäî÷íîãî ñîêà) ðàâíà 1, à ñàëüìèíà (îñíîâíîé áåëîê èç ìîëîêè ñåìãè)—ïî÷òè 12. Â èçîýëåêòðè÷åñêîé òî÷êå áåëêè íàèìåíåå óñòîé÷èâû â ðàñòâîðå è ëåãêî âûïàäàþò â îñàäîê. Èçîýëåêòðè÷åñêàÿ òî÷êà áåëêà â ñèëüíîé ñòåïåíè çàâèñèò îò ïðèñóòñòâèÿ â ðàñòâîðå èîíîâ ñîëåé; â òî æå âðåìÿ íà åå âåëè÷èíó íå âëèÿåò êîíöåíòðàöèÿ áåëêà. Â õèìèè áåëêîâ ñóùåñòâóåò ïîíÿòèå «èçîèîííàÿ òî÷êà áåëêà». Ðàñòâîð áåëêà íàçûâàåòñÿ èçîèîííûì, åñëè îí íå ñîäåðæèò íèêàêèõ äðóãèõ èîíîâ, êðîìå èîíèçèðîâàííûõ îñòàòêîâ àìèíîêèñëîò áåëêîâîé ìîëåêóëû è èîíîâ, îáðàçóþùèõñÿ ïðè äèññîöèàöèè âîäû. Äëÿ îñâîáîæäåíèÿ áåëêà îò ïîñòîðîííèõ èîíîâ îáû÷íî åãî ðàñòâîð ïðîïóñêàþò ÷åðåç êîëîíêó, íàïîëíåííóþ ñìåñüþ àíèîíî- è êàòèîíîîáìåííèêîâ. Èçîèîííîé òî÷êîé äàííîãî áåëêà ïðèíÿòî íàçûâàòü çíà÷åíèå ðÍ èçîèîííîãî ðàñòâîðà ýòîãî áåëêà: ãäå [Ð] — ìîëÿðíàÿ êîíöåíòðàöèÿ áåëêà; 2 — ñðåäíèé çàðÿä ìîëåêóëû. Ñîãëàñíî ýòîìó óðàâíåíèþ, èçîèîííàÿ òî÷êà áåëêà çàâèñèò îò åãî êîíöåíòðàöèè. Î÷åâèäíî, ïîýòîìó áåëîê, çà èñêëþ÷åíèåì ñëó÷àÿ, êîãäà ð1 ðàâíî 7, íå ìîæåò áûòü îäíîâðåìåííî èçîýëåêòðè÷åñêèì è èçîèîííûì. ÑÒÐÓÊÒÓÐÍÀß ÎÐÃÀÍÈÇÀÖÈß ÁÅËÊÎÂ Âûÿñíåíèå ñòðóêòóðíîé îðãàíèçàöèè áåëêîâ ñ÷èòàåòñÿ îäíîé èç ãëàâíûõ ïðîáëåì ñîâðåìåííîé áèîõèìèè. Îíî èìååò âàæíîå íàó÷íî-ïðàêòè÷åñêîå çíà÷åíèå äëÿ ïîíèìàíèÿ îãðîìíîãî ðàçíîîáðàçèÿ ôóíêöèé áåëêîâ, âûïîëíÿåìûõ èìè â æèâûõ îðãàíèçìàõ. Áåëêîâûå ìîëåêóëû ïðåäñòàâëÿþò ñîáîé ïðîäóêò ïîëèìåðèçàöèè 20 ðàçëè÷íûõ ìîíîìåðíûõ ìîëåêóë (àìèíîêèñëîò), ñîåäèíåííûõ íå õàîòè÷íî, à â ñòðîãîì ñîîòâåòñòâèè ñ êîäîì áåëêîâîãî ñèíòåçà (ñì. ãëàâó 14). Âîïðîñ î òîì, êàêèì îáðàçîì ñîåäèíÿþòñÿ ìåæäó ñîáîé ìíîãèå äåñÿòêè è ñîòíè àìèíîêèñëîò â áåëêîâîé ìîëåêóëå, áûë ïðåäìåòîì ïðèñòàëüíîãî âíèìàíèÿ ìíîãèõ ëàáîðàòîðèé ìèðà, çàíèìàâøèõñÿ õèìèåé áåëêà. Âïåðâûå À.ß. Äàíèëåâñêèé (1888), èçó÷àÿ áèóðåòîâóþ ðåàêöèþ, âûñêàçàë ïðåäïîëîæåíèå î ñóùåñòâîâàíèè âî âñåõ áåëêîâûõ âåùåñòâàõ îäèíàêîâûõ ãðóïï àòîìîâ è ñâÿçåé, àíàëîãè÷íûõ áèóðåòó ÊÍ2—ÑÎ—ÊÍ—ÑÎ—ÊÍ2. Òåì ñàìûì À.ß. Äàíèëåâñêèé ïåðâûé óêàçàë íà ñâÿçü —ÊÍ—ÑÎ— (ïîçäíåå ïîëó÷èâøóþ íàçâàíèå ïåïòèäíîé ñâÿçè) êàê íà íàèáîëåå âåðîÿòíûé ñïîñîá ñîåäèíåíèÿ àìèíîêèñëîò â áåëêîâîé ìîëåêóëå. Îäíàêî òîëüêî Ý. Ôèøåð (1902) ñôîðìóëèðîâàë ïîëèïåïòèäíóþ òåîðèþ ñòðîåíèÿ. Ñîãëàñíî ýòîé òåîðèè, áåëêè ïðåäñòàâëÿþò ñîáîé ñëîæíûå ïîëèïåïòèäû, â êîòîðûõ îòäåëüíûå àìèíîêèñëîòû ñâÿçàíû äðóã ñ äðóãîì ïåïòèäíûìè ñâÿçÿìè, âîçíèêàþùèìè ïðè âçàèìîäåéñòâèè à-êàðáîêñèëüíûõ ÑÎÎÍ- è à-ÌÍ2-ãðóïï àìèíîêèñëîò. Íà ïðèìåðå âçàèìîäåéñòâèÿ àëàíèíà è ãëèöèíà îáðàçîâàíèå ïåïòèäíîé ñâÿçè è äèïåïòèäà (ñ âûäåëåíèåì ìîëåêóëû âîäû) ìîæíî ïðåäñòàâèòü ñëåäóþùèì óðàâíåíèåì: ñíç-ñí—ñîîí í2ì-ñí2 ^2° ÑÍ3-ÑÍ-[ÑÎÅÌÍÜÑÍ2 1 + I —I I NN2 ÑÎÎÍ N42 ÑÎÎÍ Àëàíèí Ãëèöèí Àëàíèëãëèöèí Àíàëîãè÷íûì ñïîñîáîì ê äèïåïòèäó ìîãóò ïðèñîåäèíÿòüñÿ è äðóãèå àìèíîêèñëîòû ñ îáðàçîâàíèåì òðè-, òåòðà-, ïåíòàïåïòèäà è ò.ä. âïëîòü äî êðóïíîé ìîëåêóëû ïîëèïåïòèäà (áåëêà). Íàèìåíîâàíèå ïåïòèäîâ ñêëàäûâàåòñÿ èç íàçâàíèÿ ïåðâîé Ì-êîíöåâîé àìèíîêèñëîòû ñî ñâîáîäíîé ÌÍ2- ãðóïïîé (ñ îêîí÷àíèåì -èë, òèïè÷íûì äëÿ àöèëîâ), íàçâàíèé ïîñëåäóþùèõ àìèíîêèñëîò (òàêæå ñ îêîí÷àíèÿìè -èë) è ïîëíîãî íàçâàíèÿ Ñ-êîíöåâîé àìèíîêèñëîòû ñî ñâîáîäíîé ÑÎÎÍ-ãðóïïîé. Íàïðèìåð, ïåíòàïåïòèä èç 5 àìèíîêèñëîò ìîæåò áûòü îáîçíà÷åí ïîëíûì íàèìåíîâàíèåì: ãëèöèë- àëàíèë-ñåðèë-öèñòåèíèë-àëàíèí, èëè ñîêðàùåííî Ãëè—Àëà—Ñåð—Öèñ—Àëà . Îáðàçîâàíèå ïåïòèäíûõ ñâÿçåé, íàïðèìåð, èç òðåõ ðàçíûõ àìèíîêèñëîò ìîæåò áûòü ïðåäñòàâëåíî â âèäå ñëåäóþùåé ñõåìû: Õèìè÷åñêèé ñèíòåç ïîëèïåïòèäîâ è ñîâðåìåííûå ôèçèêî-õèìè÷åñêèå ìåòîäû èññëåäîâàíèÿ áåëêîâ ïîëíîñòüþ ïîäòâåðäèëè ñóùåñòâîâàíèå ïåïòèäíûõ ñâÿçåé â ñòðóêòóðå áåëêà. Ïîëó÷åíû ñëåäóþùèå ýêñïåðèìåíòàëüíûå äîêàçàòåëüñòâà ïîëèïåïòèäíîé òåîðèè ñòðîåíèÿ áåëêà. Â ïðèðîäíûõ áåëêàõ ñðàâíèòåëüíî ìàëî òèòðóåìûõ ñâîáîäíûõ ÑÎÎÍ- è ÌÍ2-ãðóïï, ïîñêîëüêó àáñîëþòíîå èõ áîëüøèíñòâî íàõîäèòñÿ â ñâÿçàííîì ñîñòîÿíèè, ó÷àñòâóÿ â îáðàçîâàíèè ïåïòèäíûõ ñâÿçåé; òèòðîâàíèþ äîñòóïíû â îñíîâíîì ñâîáîäíûå ÑÎÎÍ- è ÌÍ2-ãðóïïû ó Ì- è Ñ- êîíöåâûõ àìèíîêèñëîò ïåïòèäà. Â ïðîöåññå êèñëîòíîãî èëè ùåëî÷íîãî ãèäðîëèçà áåëêà îáðàçóþòñÿ ñòåõèîìåòðè÷åñêèå êîëè÷åñòâà òèòðóåìûõ ÑÎÎÍ- è ÌÍ2-ãðóïï, ÷òî ñâèäåòåëüñòâóåò î ðàñïàäå îïðåäåëåííîãî ÷èñëà ïåïòèäíûõ ñâÿçåé. Ïîä äåéñòâèåì ïðîòåîëèòè÷åñêèõ ôåðìåíòîâ (ïðîòåèíàç) áåëêè ðàñ-ùåïëÿþòñÿ íà ñòðîãî îïðåäåëåííûå ôðàãìåíòû, íàçûâàåìûå ïåïòèäàìè, ñ êîíöåâûìè àìèíîêèñëîòàìè, ñîîòâåòñòâóþùèìè èçáèðàòåëüíîñòè äåéñòâèÿ ïðîòåèíàç. Ñòðóêòóðà íåêîòîðûõ òàêèõ ôðàãìåíòîâ íåïîëíîãî ãèäðîëèçà äîêàçàíà ïîñëåäóþùèì õèìè÷åñêèì èõ ñèíòåçîì. Áèóðåòîâóþ ðåàêöèþ (ñèíå-ôèîëåòîâîå îêðàøèâàíèå â ïðèñóòñòâèè ðàñòâîðà ñóëüôàòà ìåäè â ùåëî÷íîé ñðåäå) äàþò êàê áèóðåò, ñîäåðæàùèé ïåïòèäíóþ ñâÿçü, òàê è áåëêè, ÷òî òàêæå ÿâëÿåòñÿ äîêàçàòåëüñòâîì íàëè÷èÿ â áåëêàõ àíàëîãè÷íûõ ñâÿçåé. Àíàëèç ðåíòãåíîãðàìì êðèñòàëëîâ áåëêîâ ïîäòâåðæäàåò ïîëèïåïòèä- íóþ ñòðóêòóðó áåëêîâ. Òàêèì îáðàçîì, ðåíòãåíîñòðóêòóðíûé àíàëèç ïðè ðàçðåøåíèè 0,15-0,2 íì ïîçâîëÿåò íå òîëüêî âû÷èñëèòü ìåæàòîìíûå ðàññòîÿíèÿ è ðàçìåðû âàëåíòíûõ óãëîâ ìåæäó àòîìàìè Ñ, Í, Î è Ê, íî è «óâèäåòü» êàðòèíó îáùåãî ðàñïîëîæåíèÿ àìèíîêèñëîòíûõ îñòàòêîâ â ïîëèïåïòèäíîé öåïè è ïðîñòðàíñòâåííóþ åå îðèåíòàöèþ (êîíôîðìàöèþ). Ñóùåñòâåííûì ïîäòâåðæäåíèåì ïîëèïåïòèäíîé òåîðèè ñòðîåíèÿ áåëêà ÿâëÿåòñÿ âîçìîæíîñòü ñèíòåçà ÷èñòî õèìè÷åñêèìè ìåòîäàìè ïîëèïåïòèäîâ è áåëêîâ ñ óæå èçâåñòíûì ñòðîåíèåì: èíñóëèíà - 51 àìèíîêèñëîòíûé îñòàòîê, ëèçîöèìà - 129 àìèíîêèñëîòíûõ îñòàòêîâ, ðèáîíóêëåàçû - 124 àìèíîêèñëîòíûõ îñòàòêà . Ñèíòåçèðîâàííûå áåëêè îáëàäàëè àíàëîãè÷íûìè ïðèðîäíûì áåëêàì ôèçèêî-õèìè÷åñêèìè ñâîéñòâàìè è áèîëîãè÷åñêîé àêòèâíîñòüþ. Ïîëèïåïòèäíàÿ òåîðèÿ ñòðîåíèÿ íå îòðèöàåò ñóùåñòâîâàíèÿ â ìîëåêóëå áåëêà è äðóãèõ ñâÿçåé, âêëþ÷àÿ êîâàëåíòíûå (íàïðèìåð, äèñóëüôèäíûå —8—8-ñâÿçè) è íåêîâàëåíòíûå (íàïðèìåð, âîäîðîäíûå ñâÿçè è äð.). Îíè áóäóò ðàññìîòðåíû äàëåå. Ïåïòèäíûå ñâÿçè èãðàþò èñêëþ÷èòåëüíóþ ðîëü êàê â «àðõèòåêòóðå», òàê è â ôóíêöèè áåëêîâ. Ïîýòîìó ñëåäóåò óêàçàòü íà íåêîòîðûå îñîáåííîñòè ñòðîåíèÿ ïîëèïåïòèäíîé öåïè. Âî-ïåðâûõ, ýòî ñâîåîáðàçèå ðàñïîëîæåíèÿ àòîìîâ óãëåðîäà è àçîòà, íàõîäÿùèõñÿ ïðèìåðíî â îäíîé ïëîñêîñòè, è àòîìîâ âîäîðîäà è ðàäèêàëîâ, íàïðàâëåííûõ ê ýòîé ïëîñêîñòè ïîä óãëîì 109°28'. Âî-âòîðûõ, ýòî ñâîåîáðàçèå ïåòèäíîé ñâÿçè. Ðàññòîÿíèå ìåæäó àòîìàìè Ñ è N â ïåïòèäíîé ñâÿçè (ðàâíîå 0,132 íì) ÿâëÿåòñÿ ïðîìåæóòî÷íûì ìåæäó ïðîñòîé (îðäèíàðíîé) ñâÿçüþ (ñâÿçü —Ñ—Ê—, ðàâíàÿ 147 íì) è äâîéíîé ñâÿçüþ (ñâÿçü —Ñ=Ê—, ðàâíàÿ 0,125 íì). Ýòî ñîçäàåò ïðåäïîñûëêè äëÿ îñóùåñòâëåíèÿ ïî ìåñòó äâîéíîé ñâÿçè òàóòîìåðíûõ ïåðåãðóïïèðîâîê è äëÿ îáðàçîâàíèÿ åíîëüíîé (ëàêòèìíîé) ôîðìû. Ïîñëåäíÿÿ â ñâîþ î÷åðåäü äàåò ìîëåêóëå áåëêà ðÿä ïðåèìóùåñòâ (ïîâûøåíèå ðåàêöèîííîé ñïîñîáíîñòè, âîçíèêíîâåíèå äîïîëíèòåëüíûõ âîçìîæíîñòåé âðàùåíèÿ è äð.): Ëàêòàìíàÿ (êåòîííàÿ) ôîðìà Ëàêòèìíàÿ (åíîëüíàÿ) ôîðìà Íàêîíåö, ñëåäóåò óêàçàòü íà ñâîåîáðàçèå ðàäèêàëîâ, êîòîðûå ÿâëÿþòñÿ ïîëèôóíêöèîíàëüíûìè, íåñóùèìè ñâîáîäíûå NÍ2-, ÑÎÎÍ-, ÎÍ-, 8Í- ãðóïïû è, êàê áûëî óêàçàíî, îïðåäåëÿþò ñòðóêòóðó (ïðîñòðàíñòâåííóþ) è ìíîãîîáðàçèå ôóíêöèé ìîëåêóë áåëêà. Âçàèìîäåéñòâóÿ ñ îêðóæàþùèìè ìîëåêóëàìè ðàñòâîðèòåëÿ (Í2Î), ôóíêöèîíàëüíûå ãðóïïû (â ÷àñòíîñòè, NÍ2- è ÑÎÎÍ-ãðóïïû) èîíèçèðóþòñÿ, ÷òî ïðèâîäèò ê îáðàçîâàíèþ àíèîííûõ è êàòèîííûõ öåíòðîâ áåëêîâîé ìîëåêóëû. Â çàâèñèìîñòè îò ñîîòíîøåíèÿ èîíîâ ìîëåêóëû áåëêà ïîëó÷àþò ñóììàðíûé ïîëîæèòåëüíûé (+) èëè îòðèöàòåëüíûé (—) çàðÿä ñ îïðåäåëåííûì çíà÷åíèåì èçîýëåêòðè÷åñêîé òî÷êè. Ïîëó÷åíû äîêàçàòåëüñòâà ïðåäïîëîæåíèÿ Ê. Ëèíäåðñòð¸ìà-Ëàíãà î ñóùåñòâîâàíèè 4 óðîâíåé ñòðóêòóðíîé îðãàíèçàöèè áåëêîâîé ìîëåêóëû: ïåðâè÷íîé, âòîðè÷íîé, òðåòè÷íîé è ÷åòâåðòè÷íîé ñòðóêòóðû. Òåõíèêà ñîâðåìåííîé áåëêîâîé õèìèè ðàçðàáîòàíà íàñòîëüêî õîðîøî, ÷òî ïîçâîëÿåò â ïðèíöèïå ðàñøèôðîâàòü ñòðóêòóðíóþ îðãàíèçàöèþ ëþáîãî áåëêà. Ïåðâè÷íàÿ ñòðóêòóðà áåëêà Ê íàñòîÿùåìó âðåìåíè ðàñøèôðîâàíà ïåðâè÷íàÿ ñòðóêòóðà äåñÿòêîâ òûñÿ÷ ðàçíûõ áåëêîâ, ÷òî ÿâëÿåòñÿ íåñîìíåííûì äîñòèæåíèåì áèîõèìèè. Îäíàêî 1 À12 ýòî ÷èñëî íè÷òîæíî ìàëî, åñëè ó÷åñòü, ÷òî â ïðèðîäå îêîëî 10 ðàçíîîáðàçíûõ áåëêîâ. Ïîä ïåðâè÷íîé ñòðóêòóðîé ïîäðàçóìåâàþò ïîðÿäîê, ïîñëåäîâàòåëüíîñòü ðàñïîëîæåíèÿ àìèíîêèñëîòíûõ îñòàòêîâ â ïîëèïåï- òèäíîé öåïè. Çíàÿ ïåðâè÷íóþ ñòðóêòóðó, ìåñòîïîëîæåíèå êàæäîãî îñòàòêà àìèíîêèñëîòû, ìîæíî òî÷íî íàïèñàòü ñòðóêòóðíóþ ôîðìóëó áåëêîâîé ìîëåêóëû, åñëè îíà ïðåäñòàâëåíà îäíîé ïîëèïåïòèäíîé öåïüþ. Åñëè â ñîñòàâ áåëêà âõîäèò íåñêîëüêî ïîëèïåïòèäíûõ öåïåé, îáúåäèíåííûõ â îäíó áåëêîâóþ ìîëåêóëó ïîñðåäñòâîì äèñóëüôèäíûõ ñâÿçåé è íåêîâàëåíòíûõ âçàèìîäåéñòâèé, èëè åñëè îäíà ïîëèïåïòèäíàÿ öåïü ñîäåðæèò âíóòðåííèå äèñóëüôèäíûå ñâÿçè, òî çàäà÷à îïðåäåëåíèÿ ïåðâè÷íîé ñòðóêòóðû íåñêîëüêî îñëîæíÿåòñÿ, òàê êàê íåîáõîäèìî ïðåäâàðèòåëüíîå ðàçúåäèíåíèå ýòèõ öåïåé è ñâÿçåé. Ðàçúåäèíåíèå òàêèõ ïîëèïåïòèäíûõ öåïåé ïðîèçâîäÿò ñ ïîìîùüþ äåíàòóðèðóþùèõ àãåíòîâ (ðàñòâîðû 8Ì ìî÷åâèíû èëè 6Ì ãóàíèäèíãèäðîõëîðèäà), ðàçðûâàþùèõ íåêîâàëåíòíûå ñâÿçè. Äèñóëüôèä- íûå ñâÿçè ðàçðóøàþò ïóòåì îêèñëåíèÿ èëè âîññòàíîâëåíèÿ (íàäìóðàâüèíîé êèñëîòîé èëè Ð-ìåðêàïòîýòàíîëîì ñîîòâåòñòâåííî), ïðè ýòîì îáðàçóþòñÿ ñâîáîäíûå ïîëèïåïòèäû, ñîäåðæàùèå èëè îñòàòêè öèñòåèíîâîé êèñëîòû, èëè öèñòåèíà: ì- N Äëÿ îïðåäåëåíèÿ ïåðâè÷íîé ñòðóêòóðû îòäåëüíîé, õèìè÷åñêè ãîìîãåííîé ïîëèïåïòèäíîé öåïè â ïåðâóþ î÷åðåäü ìåòîäàìè ãèäðîëèçà âûÿñíÿþò àìèíîêèñëîòíûé ñîñòàâ, òî÷íåå, ñîîòíîøåíèå êàæäîé èç 20 àìèíîêèñëîò â îáðàçöå ãîìîãåííîãî ïîëèïåïòèäà. Çàòåì ïðèñòóïàþò ê îïðåäåëåíèþ õèìè÷åñêîé ïðèðîäû êîíöåâûõ àìèíîêèñëîò ïîëèïåïòèäíîé öåïè, ñîäåðæàùåé îäíó ñâîáîäíóþ NÍ2-ãðóïïó è îäíó ñâîáîäíóþ ÑÎÎÍ-ãðóïïó. Ìåòîäû îïðåäåëåíèÿ Û-êîíöåâîé àìèíîêèñëîòû Äëÿ îïðåäåëåíèÿ ïðèðîäû Ê-êîíöåâîé àìèíîêèñëîòû ïðåäëîæåí ðÿä ìåòîäîâ, â ÷àñòíîñòè ìåòîä Ñýíäæåðà (Ð. 8àï§åã), îñíîâàííûé íà ðåàêöèè àðèëèðîâàíèÿ ïîëèïåïòèäà 2,4-äèíèòðîôòîðáåíçîëîì (ÄÍÔÁ), ÷òî ïðèâîäèò ê îáðàçîâàíèþ îêðàøåííîãî â æåëòûé öâåò 2,4-äèíèòðîôåíèëüíîãî ïðîèçâîäíîãî Ê-êîíöåâîé àìèíîêèñëîòû . Ðàñòâîð ïîëèïåïòèäà îáðàáàòûâàþò ÄÍÔÁ, êîòîðûé âçàèìîäåéñòâóåò ñî ñâîáîäíîé ÊÍ2-ãðóïïîé Ê-êîíöåâîé àìèíîêèñëîòû ïåïòèäà. N02 -ÑÍ-ÑÎ-ÌÍ-ÑÍ-ÑÎ—ìí-ñí-ñîîí Ðï ê2 Ïîëèïåïòèä(áåëîê) V N02 "N02 íãä-ñí-ñî—ìí-ñí-ñî ÌÍ-ÑÍ-ÑÎÎÍ I I Ê2 Êï Ãèäðîëèç + Ñâîáîäíûå àìèíîêèñëîòû , N02 ÍÃ^-ÑÍ-ÑÎÎÍ I Ê\| Äèíèòðîôåíèëàìèíîêèñëîòà Ïîñëå êèñëîòíîãî ãèäðîëèçà ïðîäóêòà ðåàêöèè—äèíèòðîôåíèëïåïòèäà òîëüêî îäíà Ê-êîíöåâàÿ àìèíîêèñëîòà îêàçûâàåòñÿ ñâÿçàííîé ñ ðåàêòèâîì â âèäå 2,4-äèíèòðîôåíèëàìèíîêèñëîòû (ñòàáèëüíîé ïðè ãèäðîëèçå). Â îòëè÷èå îò äðóãèõ îáðàçîâàâøèõñÿ ïðè ãèäðîëèçå ïîëèïåïòèäà ñâîáîäíûõ àìèíîêèñëîò îíà æåëòîãî öâåòà. Åå èäåíòèôèöèðóþò ìåòîäîì õðîìàòîãðàôèè. Äëÿ îïðåäåëåíèÿ Ê-êîíöåâîé àìèíîêèñëîòû çíà÷èòåëüíî áîëåå øèðîêî ïðèìåíÿåòñÿ ôåíè ëòèîãèäàíòîèíîâûé ìåòîä Ýäìàíà áëàãîäàðÿ ñâîåé âûñîêîé ÷óâñòâèòåëüíîñòè è âîçìîæíîñòè ìíîãîêðàòíîãî èñïîëüçîâàíèÿ â îäíîé è òîé æå ïðîáå. Ôåíèëèçîòèîöèàíàò ðåàãèðóåò ñî ñâîáîäíîé à-ÊÈ2-ãðóïïîé Ê-êîíöåâîé àìèíîêèñëîòû ïîëèïåïòèäà ñ îáðàçîâàíèåì ôåíèëòèîêàðáàìîèëïåïòèäà. N=0=3 + ÍãÌ-ÑÍ-ÑÎ^Í-ÑÍ-ÑÎ NN-04-0004 1 I Ôåíèëèçîòèîöèàíàò I Ï0ëèïåïòèä N4 ÑÎ—N4—ÎÍ—ÑÎ N4—04-0004 I1 I Ã,^Ñ\ /ÑÍ-Ð, Ð2 Ðï N4 ÍÑ1 õ N Ñ=0 >.V Í2N-ÑÍ-Ñ0 N4-04-0004 I \| ^Ñ\ /ÑÍ-«1 ð2 Ðë “ N4 ôåíèëïÒ1èñýãèä;àíòîèíîâîå Ïîëèïåïòèä, óêîðî÷åííûé ïðîèçâîäíîå íà îäíó àìèíîêèñëîòó ^êîíöåâîé ÀÌÊ ’ ’ Îáðàáîòêà ïðîäóêòà ðåàêöèè êèñëîòîé ïðèâîäèò ê öèêëèçàöèè è îñâîáîæäåíèþ ôåíèëòèîãèäàíòîèíà ^êîíöåâîé àìèíîêèñëîòû, ïðèðîäó êîòîðîãî óñòàíàâëèâàþò õðîìàòîãðàôè÷åñêè. Óêîðî÷åííûé íà îäíó àìèíîêèñëîòó ïîëèïåïòèä ïîäâåðãàþò äàëüíåéøåìó àíàëèçó. Ýòó ïðîöåäóðó ñòóïåí÷àòîãî ðàñùåïëåíèÿ ïåïòèäà ñ Îêîíöà ìîæíî ïîâòîðÿòü ìíîãîêðàòíî, èäåíòèôèöèðóÿ ïîñëåäîâàòåëüíî îäíó àìèíîêèñëîòó çà äðóãîé. Ìåòîä Ýäìàíà èñïîëüçóåòñÿ â êà÷åñòâå õèìè÷åñêîé îñíîâû äëÿ îïðåäåëåíèÿ ïåðâè÷íîé ñòðóêòóðû áåëêîâ è ïåïòèäîâ. Îí ðåàëèçîâàí â ñïåöèàëüíîì ïðèáîðå — ñåêâåíàòîðå (îò àíãë. çåäèåïñå — ïîñëåäîâàòåëüíîñòü), ðàáîòàþùåì â àâòîìàòè÷åñêîì ðåæèìå è ïîçâîëÿþùåì îïðåäåëèòü ïîñëåäîâàòåëüíîñòü àìèíîêèñëîò ñ Îêîíöà ïåïòèäà äî 50—60 àìèíîêèñëîòíûõ îñòàòêîâ. Êðîìå ýòèõ ðåàêòèâîâ, äëÿ îïðåäåëåíèÿ ÷åðåäîâàíèÿ àìèíîêèñëîò èñïîëüçóþò öèàíàò êàëèÿ, 1-äèìåòèëàìèíîíàôòèë-5-ñóëüôîíèëõëîðèä—äàí- ñèëõëîðèä è äàáñèëõëîðèä. Äëÿ ýòèõ æå öåëåé èíîãäà ïðèìåíÿþò ôåðìåíòû ýêçîïåïòèäàçû, â ÷àñòíîñòè àëàíèí- è ëåéöèíàìèíîïåïòèäàçó. Ýòè ôåðìåíòû ðàçðûâàþò ïåïòèäíûå ñâÿçè ñ òîãî êîíöà ïîëèïåïòèäà, ãäå èìååòñÿ ñâîáîäíàÿ NÍ2-ãðóïïà, îñâîáîæäàÿ ^êîíöåâóþ àìèíîêèñëîòó (ìåõàíèçì äåéñòâèÿ ýêçîïåïòèäàç ñì. â ãëàâå 12). Ìåòîäû îïðåäåëåíèÿ Ñ-êîíöåâîé àìèíîêèñëîòû Äëÿ îïðåäåëåíèÿ ïðèðîäû Ñ-êîíöåâîé àìèíîêèñëîòû ÷àñòî èñïîëüçóþò ôåðìåíòàòèâíûå ìåòîäû. Îáðàáîòêà ïîëèïåïòèäà êàðáîêñèïåïòèäàçîé, êîòîðàÿ ðàçðûâàåò ïåïòèäíóþ ñâÿçü ñ òîãî êîíöà ïåïòèäà, ãäå ñîäåðæèòñÿ ñâîáîäíàÿ ÑÎÎÍ-ãðóïïà, ïðèâîäèò ê îñâîáîæäåíèþ Ñ-êîíöåâîé àìèíîêèñëîòû, ïðèðîäà êîòîðîé ìîæåò áûòü èäåíòèôèöèðîâàíà ìåòîäîì õðîìàòîãðàôèè. Ïðåäëîæåí òàêæå õèìè÷åñêèé ìåòîä Àêàáîðè (8. ÀêàÚîï), êîòîðûé îñíîâàí íà ãèäðàçèíîëèçå ïîëèïåïòèäà: í2ì-ñí-ñî<-ìí-ñí-ñî)ï-(\ò-ñí-ñîîí+<ï+\|)í2ã-ãø2 I I ð, ð 0Íä Ãèäðàçèí —> (ï+1)Í!Ì-ÑÍ-Ñ0 + ÍýÌ-ÑÍ-ÑÎÎÍ NN-N42 ÑÍç Àìèíîàöèëãèäðàçèíû Àëàíèí Ãèäðàçèí, âûçûâàÿ ðàñïàä ÷óâñòâèòåëüíûõ ê íåìó ïåïòèäíûõ ñâÿçåé ïîëèïåïòèäà, ðåàãèðóåò ñî âñåìè àìèíîêèñëîòàìè, çà èñêëþ÷åíèåì Ñ- êîíöåâîé àìèíîêèñëîòû, ïîñêîëüêó åå êàðáîêñèëüíàÿ ãðóïïà íå ó÷àñòâóåò â îáðàçîâàíèè ïåïòèäíîé ñâÿçè. Ïðè ýòîì îáðàçóåòñÿ ñìåñü àìèíîàöèë- ãèäðàçèíîâ è ñâîáîäíîé Ñ-êîíöåâîé àìèíîêèñëîòû. Ïîñëåäíþþ ïîñëå îáðàáîòêè âñåé ñìåñè ÄÍÔÁ îòäåëÿþò è èäåíòèôèöèðóþò õðîìàòîãðàôè÷åñêè, äëÿ ÷åãî îáðàçîâàâøèåñÿ äèíèòðîôåíèëïðîèçâîäíûå àìèíîàöèë- ãèäðàçèíîâ ïðåäâàðèòåëüíî ýêñòðàãèðóþò óêñóñíî-ýòèëîâûì ýôèðîì. Ñ-êîíöåâóþ àìèíîêèñëîòó èäåíòèôèöèðóþò òàêæå ïóòåì îáðàáîòêè ïîëèïåïòèäà âîññòàíàâëèâàþùèì àãåíòîì, íàïðèìåð áîðãèäðèäîì íàòðèÿ. Â ïðîñòåéøåé ôîðìå ýòó ïðîöåäóðó ìîæíî ïðåäñòàâèòü â ñëåäóþùåì âèäå: Í2Ì-ÑÍ-ÑÎ-(ÌÍ-ÑÍ-ÑÎ)ï-ÌÍ-ÑÍ-ÑÎ ÌÍ-ÑÍ-ÑÎÎÍ ^ I I ÛàÂÈ, Í2Ì-ÑÍ-ÑÎ-(ÌÍ-ÑÍ-ÑÎ>ï-ÌÍ-ÑÍ-ÑÎ ÌÍ-ÑÍ-ÑÍãÎÍ I 6Ì ÍÑ1, 105°Ñ, 24÷ Í21×-ÑÍ-ÑÎÎÍ (Í2Ì-ÑÍ-ÑÎÎÍ)ï Í2Ì-ÑÍ-ÑÎÎÍ í2«-ÑÍ-ÑÍîÎÍ +1 +1 + I Ê\| Ê* Ê ð Ñâîáîäíûå àìèíîêèñëîòû Âèäíî, ÷òî â óêàçàííûõ óñëîâèÿõ òîëüêî îäíà, à èìåííî Ñ-êîíöåâàÿ, àìèíîêèñëîòà áóäåò ïðåâðàùàòüñÿ â à-àìèíîñïèðò, ëåãêî èäåíòèôèöèðóåìûé ìåòîäîì õðîìàòîãðàôèè. Òàêèì îáðàçîì, ïðè ïîìîùè óêàçàííûõ ìåòîäîâ îïðåäåëÿþò ïðèðîäó N è Ñ-êîíöåâûõ àìèíîêèñëîò. Ñëåäóþùèé ýòàï ðàáîòû ñâÿçàí ñ îïðåäåëåíèåì ÷åðåäîâàíèÿ (ïîñëåäî-âàòåëüíîñòè) àìèíîêèñëîò âíóòðè ïîëèïåïòèäíîé öåïè. Äëÿ ýòîãî ñíà÷àëà ïðîâîäÿò èçáèðàòåëüíûé, ÷àñòè÷íûé (õèìè÷åñêèé è ôåðìåíòàòèâíûé), ãèäðîëèç ïîëèïåïòèäíîé öåïè íà êîðîòêèå ïåïòèäíûå ôðàãìåíòû, ïîñëåäîâàòåëüíîñòü àìèíîêèñëîò â êîòîðûõ ìîæåò áûòü òî÷íî îïðåäåëåíà îïèñàííûìè ðàíåå ìåòîäàìè. Õèìè÷åñêèå ìåòîäû èçáèðàòåëüíîãî è íåïîëíîãî ãèäðîëèçà îñíîâàíû íà ïðèìåíåíèè òàêèõ õèìè÷åñêèõ ðåàêòèâîâ, êîòîðûå âûçûâàþò ñåëåêòèâíûé, âûñîêîñïåöèôè÷åñêèé ðàçðûâ ïåïòèäíûõ ñâÿçåé, îáðàçîâàííûõ îïðåäåëåííûìè àìèíîêèñëîòàìè, îñòàâëÿÿ íåçàòðîíóòûìè îñòàëüíûå ïåïòèäíûå ñâÿçè. Ê ýòèì èçáèðàòåëüíî ãèäðîëèçóþùèì âåùåñòâàì îòíîñÿòñÿ öèàíîãåíáðîìèä, ˆN61 (ïî îñòàòêàì ìåòèîíèíà), ãèäðîêñèëàìèí (ïî ñâÿçÿì ìåæäó îñòàòêàìè àñïàðàãèíîâîé êèñëîòû è ãëèöèíà), ^áðîìñóêöèíà- ìèä (ïî îñòàòêàì òðèïòîôàíà). Ìåòèîíèíà â ñîñòàâå áåëêîâ ñîäåðæèòñÿ îáû÷íî ìåíüøå, ÷åì äðóãèõ àìèíîêèñëîò, ïîýòîìó îáðàáîòêà ÑÊÂã ïðåäïî÷òèòåëüíåå, òàê êàê ïðè ýòîì îáðàçóåòñÿ íåáîëüøîå ÷èñëî ïåïòèäîâ, ïåðâè÷íóþ ñòðóêòóðó êîòîðûõ îïðåäåëÿþò ñ ïîìîùüþ ðàññìîòðåííûõ ðàíåå ìåòîäîâ, âñÿêèé ðàç íà÷èíàÿ ñ îïðåäåëåíèÿ ïðèðîäû Ê- è Ñ-êîíöåâûõ àìèíîêèñëîò. Ôåðìåíòàòèâíûå ìåòîäû ãèäðîëèçà îñíîâàíû íà èçáèðàòåëüíîñòè äåéñòâèÿ ïðîòåîëèòè÷åñêèõ (âûçûâàþùèõ ðàñïàä áåëêîâ) ôåðìåíòîâ, ðàñùåïëÿþùèõ ïåïòèäíûå ñâÿçè, îáðàçîâàííûå îïðåäåëåííûìè àìèíîêèñëîòàìè. Â ÷àñòíîñòè, ïåïñèí óñêîðÿåò ãèäðîëèç ñâÿçåé, îáðàçîâàííûõ îñòàòêàìè ôåíèëàëàíèíà, òèðîçèíà è ãëóòàìèíîâîé êèñëîòû, òðèïñèí — àðãèíèíà è ëèçèíà, õèìîòðèïñèí—òðèïòîôàíà, òèðîçèíà è ôåíèëàëàíèíà. Ðÿä äðóãèõ ôåðìåíòîâ, íàïðèìåð ïàïàèí, ñóáòèëèçèí, ïðîíàçà è äðóãèå áàêòåðèàëüíûå ïðîòåèíàçû, òàêæå èñïîëüçóåòñÿ äëÿ íåïîëíîãî ãèäðîëèçà áåëêîâ. Â ðåçóëüòàòå ïîëèïåïòèäíàÿ öåïü ðàñùåïëÿåòñÿ íà ìåëêèå ïåïòèäû, ñîäåðæàùèå èíîãäà âñåãî íåñêîëüêî àìèíîêèñëîò, êîòîðûå îòäåëÿþò äðóã îò äðóãà ñî÷åòàííûìè ýëåêòðîôîðåòè÷åñêèìè è õðîìàòîãðàôè÷åñêèìè ìåòîäàìè, ïîëó÷àÿ ñâîåîáðàçíûå ïåïòèäíûå êàðòû. Äàëåå îïðåäåëÿþò ÷åðåäîâàíèå àìèíîêèñëîò â êàæäîì èíäèâèäóàëüíîì ïåïòèäå. Çàâåðøàåòñÿ ðàáîòà âîññîçäàíèåì ïåðâè÷íîé ñòðóêòóðû ïîëíîé ïîëèïåïòèäíîé öåïè íà îñíîâàíèè îïðåäåëåíèÿ ïîñëåäîâàòåëüíîñòè àìèíîêèñëîò â îòäåëüíûõ ïåïòèäàõ. Ìåòîä ñîñòàâëåíèÿ ïåïòèäíûõ êàðò, ïîëó÷èâøèé îáðàçíîå íàçâàíèå «ìåòîä îòïå÷àòêîâ ïàëüöåâ», èñïîëüçóåòñÿ ïðè îïðåäåëåíèè ñõîäñòâà èëè ðàçëè÷èÿ ãîìîëîãè÷íûõ áåëêîâ ïî ïåðâè÷íîé ñòðóêòóðå. Áåëîê èíêóáèðóþò ñ êàêèì-ëèáî ïðîòåîëèòè÷åñêèì ôåðìåíòîì. ×àñòî ïîðöèè áåëêà èíêóáèðóþò êàê ñ ïåïñèíîì, òàê è ñ òðèïñèíîì. Ïðè ýòîì âñëåäñòâèå ãèäðîëèçà ñòðîãî îïðåäåëåííûõ ïåïòèäíûõ ñâÿçåé îáðàçóåòñÿ ñìåñü êîðîòêèõ ïåïòèäîâ, ëåãêî ðàçäåëÿåìûõ ñ ïîìîùüþ õðîìàòîãðàôèè â îäíîì íàïðàâëåíèè è ýëåêòðîôîðåçà — â äðóãîì, ïîä óãëîì 90° îò ïåðâîãî (ïåïòèäíàÿ êàðòà). Äàëüíåéøèå çàäà÷è — óñòàíîâëåíèå ïîñëåäîâàòåëüíîñòè ðàñïîëîæåíèÿ àìèíîêèñëîò â êàæäîì èç âûäåëåííûõ ïåïòèäîâ (ôåíèëòèîãèäàíòîèíîâûì èëè äðóãèìè ìåòîäàìè), ñîïîñòàâëåíèå ïîëó÷åííûõ äàííûõ è óñòàíîâëåíèå ïåðâè÷íîé ñòðóêòóðû âñåé ìîëåêóëû. Âîçìîæíîñòü ïðèìåíåíèÿ ðåíòãåíîñòðóêòóðíîãî àíàëèçà äëÿ îïðåäåëåíèÿ ïîñëåäîâàòåëüíîñòè àìèíîêèñëîò â áåëêîâîé ìîëåêóëå áûëà ðàññìîòðåíà ðàíåå. Ñëåäóåò îòìåòèòü ñîâåðøåííî íîâûé ïîäõîä ê ðåøåíèþ ýòîé âàæíîé ïðîáëåìû — îïðåäåëåíèå ïîñëåäîâàòåëüíîñòè àìèíîêèñëîò â áåëêîâîé ìîëåêóëå ñ èñïîëüçîâàíèåì äàííûõ î êîìïëåìåíòàðíîé íóêëåîòèäíîé ïîñëåäîâàòåëüíîñòè ÄÍÊ. Ýòîìó ñïîñîáñòâóþò êàê ìåòîäû áûñòðîãî ñåêâåíèðîâàíèÿ ÄÍÊ, òàê è òåõíèêà èçîëèðîâàíèÿ è äîñòóïíîñòè ñàìîãî ãåíà . Â íàñòîÿùåå âðåìÿ âûÿñíåíèå ïåðâè÷íîé ñòðóêòóðû áåëêîâ ÿâëÿåòñÿ âîïðîñîì âðåìåíè è òåõíè÷åñêîãî îñíàùåíèÿ ëàáîðàòîðèé. Ïîëíîñòüþ âûÿñíåíà ïåðâè÷íàÿ ñòðóêòóðà ìíîãèõ ïðèðîäíûõ áåëêîâ è ïðåæäå âñåãî èíñóëèíà, ñîäåðæàùåãî 51 àìèíîêèñëîòíûé îñòàòîê [Ñýíäæåð Ô., 1954]. Áîëåå êðóïíûì áåëêîì ñ âûÿñíåííîé ïåðâè÷íîé ñòðóêòóðîé îêàçàëñÿ èììóíîãëîáóëèí, â ÷åòûðåõ ïîëèïåïòèäíûõ öåïÿõ êîòîðîãî íàñ÷èòûâàåòñÿ 1300 àìèíîêèñëîòíûõ îñòàòêîâ. Çà ýòó ðàáîòó Äæ. Ýäåëüìàí è Ð. Ïîðòåð áûëè óäîñòîåíû Íîáåëåâñêîé ïðåìèè (1972). Ðèñ. 1.14. Ñòðóêòóðà ïðîèíñóëèíà. Ðàñøèôðîâàíû ïåðâè÷íûå ñòðóêòóðû ìèîãëîáèíà ÷åëîâåêà (153 àìèíîêèñëîòíûõ îñòàòêà), à-öåïè (141) è Ð-öåïè (146) ãåìîãëîáèíà ÷åëîâåêà, öèòîõðîìà Ñ èç ñåðäå÷íîé ìûøöû ÷åëîâåêà (104), ëèçîöèìà ìîëîêà ÷åëîâåêà (130), õèìîòðèïñèíîãåíà áûêà (245) è ìíîãèõ äðóãèõ áåëêîâ, â òîì ÷èñëå ôåðìåíòîâ è òîêñèíîâ. Íà ðèñ. 1.14 ïðåäñòàâëåíà ïîñëåäîâàòåëüíîñòü àìèíîêèñëîòíûõ îñòàòêîâ ïðîèíñóëèíà. Âèäíî, ÷òî ìîëåêóëà èíñóëèíà (âûäåëåíà òåìíûìè êðóæêàìè), ñîñòîÿùàÿ èç äâóõ öåïåé (À—21 è Â — 30 àìèíîêèñëîòíûõ îñòàòêîâ), îáðàçóåòñÿ èç ñâîåãî ïðåäøåñòâåííèêà — ïðîèíñóëèíà (84 àìèíîêèñëîòíûõ îñòàòêà), ïðåäñòàâëåííîãî îäíîé ïîëèïåï- òèäíîé öåïüþ, ïîñëå îòùåïëåíèÿ îò íåãî ïåïòèäà, ñîñòîÿùåãî èç 33 àìèíîêèñëîòíûõ îñòàòêîâ. Ñòðîåíèå ìîëåêóëû èíñóëèíà (51 àìèíîêèñëîòíûé îñòàòîê) ñõåìàòè÷åñêè ìîæíî ïðåäñòàâèòü ñëåäóþùèì îáðàçîì: À-öåïü (21) Â-öåïü (30) Ìåæäó öåïÿìè À è Â è âíóòðè À-öåïè èíñóëèíà îáðàçóþòñÿ äèñóëü- ôèäíûå (—8—8—) ñâÿçè. Âûÿñíåíà ïåðâè÷íàÿ ñòðóêòóðà áîëåå 18 èíñóëè- íîâ, âûäåëåííûõ èç ðàçíûõ èñòî÷íèêîâ. Áëèçêèìè ïî ïåðâè÷íîé ñòðóêòóðå îêàçàëèñü èíñóëèíû èç ïîäæåëóäî÷íîé æåëåçû ÷åëîâåêà, ñâèíüè è êàøàëîòà. Åäèíñòâåííûì îòëè÷èåì èíñóëèíà ÷åëîâåêà ÿâëÿåòñÿ íàõîæäåíèå òðåîíèíà â ïîëîæåíèè 30 Â-öåïè âìåñòî àëàíèíà. Âòîðûì áåëêîì, ïåðâè÷íàÿ ñòðóêòóðà êîòîðîãî ðàñøèôðîâàíà Ñ. Ìóðîì è Ó. Ñòåéíîì, ÿâëÿåòñÿ ðèáîíóêëåàçà (ðèñ. 1.15) èç ïîäæåëóäî÷íîé æåëåçû, êàòàëèçèðóþùàÿ ðàñùåïëåíèå ÐÍÊ. Ôåðìåíò ñîñòîèò èç 124 àìèíîêèñëîòíûõ îñòàòêîâ ñ Ì-êîíöåâûì ëèçèíîì è Ñ-êîíöåâûì âàëèíîì, ìåæäó îñòàòêàìè öèñòåèíà îáðàçóþòñÿ äèñóëüôèäíûå (—8—8—) ñâÿçè â 4 ó÷àñòêàõ. Ïîëíîñòüþ ðàñøèôðîâàíà ïîñëåäîâàòåëüíîñòü àìèíîêèñëîò ïîëèïåï- òèäíîé öåïè ôåðìåíòà ëèçîöèìà, èìåþùåãî âàæíîå çàùèòíîå è ìåäèöèíñêîå çíà÷åíèå, òàê êàê îí âûçûâàåò ëèçèñ ðÿäà áàêòåðèé, ðàñùåïëÿÿ îñíîâíîå âåùåñòâî èõ êëåòî÷íîé îáîëî÷êè. Ëèçîöèì áåëêà êóðèíîãî ÿéöà ñîäåðæèò 129 àìèíîêèñëîò (ðèñ. 1.16) ñ Ì-êîíöåâûì ëèçèíîì è Ñ-êîíöåâûì ëåéöèíîì. Îòå÷åñòâåííûìè èññëåäîâàòåëÿìè óñòàíîâëåíà ïåðâè÷íàÿ ñòðóêòóðà ìíîãèõ áåëêîâ è ïîëèïåïòèäîâ, â òîì ÷èñëå êðóïíîãî áåëêà ÐÍÊ-ïîëèìåðà- çû (â ÷àñòíîñòè, ïîñëåäîâàòåëüíîñòè åå â- è ð'-ñóáúåäèíèö, 1342 è 1407 50 Ðèñ. 1.15. Ïåðâè÷íàÿ ñòðóêòóðà ÐÍÊàçû. Öâåòîì âûäåëåíû ÷åòûðå äèñóëüôèäíûå ñâÿçè. Ðèñ. 1.16. Ïåðâè÷íàÿ ñòðóêòóðà ïîëèïåïòèäíîé öåïè ëèçîöèìà (ñõåìà). àìèíîêèñëîòíûõ îñòàòêîâ ñîîòâåòñòâåííî ôàêòîðà ýëîíãàöèè Î èç Å.ñîÍ (701 àìèíîêèñëîòà) (Þ.À. Îâ÷èííèêîâ è äð.), ôåðìåíòà àñïàðòàòàìèíî- òðàíñôåðàçû, ñîñòîÿùåé èç 412 àìèíîêèñëîòíûõ îñòàòêîâ (À.Å. Áðàóí- øòåéí, Þ.À. Îâ÷èííèêîâ è äð.), ëåããåìîãëîáèíà, áåëêà Ü25 èç ðèáîñîì Å.ñîÍ, íåéðîòîêñèíîâ èç ÿäà êîáðû (Þ.À. Îâ÷èííèêîâ è äð.), ïåïñèíîãåíà è ïåïñèíà (Â.Ì. Ñòåïàíîâ è äð.), Ü-ëèïîòðîïèíà è ëàêòîãåííîãî ãîðìîíà áûêà (Í.À. Þäàåâ, Þ.À. Ïàíêîâ) è äð. Èññëåäîâàíèÿ ïåðâè÷íîé ñòðóêòóðû à- è ð-öåïåé ãåìîãëîáèíà ñïîñîáñòâîâàëè âûÿñíåíèþ ñòðóêòóðû íåîáû÷íûõ, òàê íàçûâàåìûõ àíîìàëüíûõ, ãåìîãëîáèíîâ, âñòðå÷àþùèõñÿ â êðîâè áîëüíûõ ãåìîãëîáèíîïàòèÿìè. Èíîãäà ðàçâèòèå áîëåçíè, êàê è èçìåíåíèå ïðîñòðàíñòâåííîé ñòðóêòóðû ãåìîãëîáèíà ÷åëîâåêà, îáóñëîâëåíî çàìåíîé ëèøü îäíîé êàêîé-ëèáî àìèíîêèñëîòû â ñòðóêòóðå ð-öåïåé (ðåæå à-öåïåé) ãåìîãëîáèíà (ñì. ãëàâó 2). Àíàëèç äàííûõ î ïåðâè÷íîé ñòðóêòóðå áåëêîâ ïîçâîëÿåò ñäåëàòü ñëåäóþùèå îáùèå âûâîäû. Ïåðâè÷íàÿ ñòðóêòóðà áåëêîâ óíèêàëüíà è äåòåðìèíèðîâàíà ãåíåòè÷åñêè. Êàæäûé èíäèâèäóàëüíûé ãîìîãåííûé áåëîê õàðàêòåðèçóåòñÿ óíèêàëüíîé ïîñëåäîâàòåëüíîñòüþ àìèíîêèñëîò: ÷àñòîòà çàìåíû àìèíîêèñëîò ïðèâîäèò íå òîëüêî ê ñòðóêòóðíûì ïåðåñòðîéêàì, íî è ê èçìåíåíèÿì ôèçèêî-õèìè÷åñêèõ ñâîéñòâ è áèîëîãè÷åñêèõ ôóíêöèé. Ñòàáèëüíîñòü ïåðâè÷íîé ñòðóêòóðû îáåñïå÷èâàåòñÿ â îñíîâíîì ãëàâ-íîâàëåíòíûìè ïåïòèäíûìè ñâÿçÿìè; âîçìîæíî ó÷àñòèå íåáîëüøîãî ÷èñëà äèñóëüôèäíûõ ñâÿçåé. Â ïîëèïåïòèäíîé öåïè ìîãóò áûòü îáíàðóæåíû ðàçíîîáðàçíûå êîìáèíàöèè àìèíîêèñëîò; â ïîëèïåïòèäàõ îòíîñèòåëüíî ðåäêè ïîâòîðÿþùèåñÿ ïîñëåäîâàòåëüíîñòè. Â íåêîòîðûõ ôåðìåíòàõ, îáëàäàþùèõ áëèçêèìè êàòàëèòè÷åñêèìè ñâîéñòâàìè, âñòðå÷àþòñÿ èäåíòè÷íûå ïåïòèäíûå ñòðóêòóðû, ñîäåðæàùèå íåèçìåííûå (èíâàðèàíòíûå) ó÷àñòêè è âàðèàáåëüíûå ïîñëåäîâàòåëüíîñòè àìèíîêèñëîò, îñîáåííî â îáëàñòÿõ èõ àêòèâíûõ öåíòðîâ. Ýòîò ïðèíöèï ñòðóêòóðíîãî ïîäîáèÿ íàèáîëåå òèïè÷åí äëÿ ðÿäà ïðîòåîëèòè÷åñêèõ ôåðìåíòîâ: òðèïñèíà, õèìîòðèïñèíà è äð. (ñì. ãëàâó 4). Â ïåðâè÷íîé ñòðóêòóðå ïîëèïåïòèäíîé öåïè äåòåðìèíèðîâàíû âòîðè÷íàÿ, òðåòè÷íàÿ è ÷åòâåðòè÷íàÿ ñòðóêòóðû áåëêîâîé ìîëåêóëû, îïðåäåëÿþùèå åå îáùóþ ïðîñòðàíñòâåííóþ êîíôîðìàöèþ. Âòîðè÷íàÿ ñòðóêòóðà áåëêà Ðåíòãåíîñòðóêòóðíàÿ êðèñòàëëîãðàôèÿ ðåøàåò äâå ãëàâíûå ïðîáëåìû áåëêîâîé õèìèè: çàêîíîìåðíîñòè ÷åðåäîâàíèÿ ïîñëåäîâàòåëüíîñòè àìèíîêèñëîòíûõ îñòàòêîâ â ïîëèïåïòèäíîé öåïè è çàêîíîìåðíîñòè êîíôèãóðàöèè áåëêîâîé ìîëåêóëû. Ïåðâûå ðåíòãåíîãðàììû áåëêîâ, ïîëó÷åííûå åùå â 30-õ ãîäàõ Ó. Àñòáþ- ðè, à çàòåì Ë. Ïîëèíãîì è Ð. Êîðè, ïîçâîëèëè óñòàíîâèòü íàëè÷èå â áåëêàõ íàðÿäó ñ ëèíåéíîé ïîëèïåïòèäíîé öåïüþ ó÷àñòêîâ, îïðåäåëåííûì îáðàçîì ñêðó÷åííûõ. Ïîä âòîðè÷íîé ñòðóêòóðîé áåëêà ïîäðàçóìåâàþò êîíôèãóðàöèþ ïîëè- ïåïòèäíîé öåïè, ò. å. ñïîñîá ñâåðòûâàíèÿ, ñêðó÷èâàíèÿ (ñêëàäûâàíèå, óïàêîâêà) ïîëèïåïòèäíîé öåïè â ñïèðàëüíóþ èëè êàêóþ-ëèáî äðóãóþ êîíôîðìàöèþ. Ïðîöåññ ýòîò ïðîòåêàåò íå õàîòè÷íî, à â ñîîòâåòñòâèè ñ ïðîãðàììîé, çàëîæåííîé â ïåðâè÷íîé ñòðóêòóðå. Ïîäðîáíî èçó÷åíû äâå îñíîâíûå êîíôèãóðàöèè ïîëèïåïòèäíûõ öåïåé, îòâå÷àþùèõ ñòðóêòóðíûì òðåáîâàíèÿì è ýêñïåðèìåíòàëüíûì äàííûì: à-ñïèðàëè è â-ñòðóêòóðû. Áëàãîäàðÿ èññëåäîâàíèÿì Ë. Ïîëèíãà íàèáîëåå âåðîÿòíûì òèïîì ñòðîåíèÿ ãëîáóëÿðíûõ áåëêîâ ïðèíÿòî ñ÷èòàòü à-ñïèðàëü (ðèñ. 1.17). Çàêðó÷èâàíèå ïîëèïåïòèäíîé öåïè ïðîèñõîäèò ïî ÷àñîâîé ñòðåëêå (ïðàâûé õîä ñïèðàëè), ÷òî îáóñëîâëåíî Ü-àìèíîêèñëîòíûì ñîñòàâîì ïðèðîäíûõ áåëêîâ. Äâèæóùåé ñèëîé â âîçíèêíîâåíèè à-ñïèðàëåé (òàê æå êàê è â-ñòðóêòóð) ÿâëÿåòñÿ ñïîñîáíîñòü àìèíîêèñëîò ê îáðàçîâàíèþ âîäîðîäíûõ ñâÿçåé. Â ñòðóêòóðå à-ñïèðàëåé îòêðûò ðÿä çàêîíîìåðíîñòåé. Íà êàæäûé âèòîê (øàã) ñïèðàëè ïðèõîäèòñÿ 3,6 àìèíîêèñëîòíûõ îñòàòêà. Øàã ñïèðàëè (ðàññòîÿíèå âäîëü îñè) ðàâåí 0,54 íì íà âèòîê, à íà îäèí àìèíîêèñëîòíûé îñòàòîê ïðèõîäèòñÿ 0,15 íì. Óãîë ïîäúåìà ñïèðàëè 26°, ÷åðåç 5 âèòêîâ ñïèðàëè (18 àìèíîêèñëîòíûõ îñòàòêîâ) ñòðóêòóðíàÿ êîíôèãóðàöèÿ ïîëèïåï- òèäíîé öåïè ïîâòîðÿåòñÿ. Ýòî îçíà÷àåò, ÷òî ïåðèîä ïîâòîðÿåìîñòè (èëè èäåíòè÷íîñòè) à-ñïèðàëüíîé ñòðóêòóðû ñîñòàâëÿåò 2,7 íì. Äëÿ êàæäîãî áåëêà õàðàêòåðíà îïðåäåëåííàÿ ñòåïåíü ñïèðàëèçàöèè åãî ïîëèïåïòèäíîé öåïè. Ñòåïåíü ñïèðàëèçàöèè óñòàíàâëèâàþò ïóòåì èçìåðåíèÿ óäåëüíîãî âðàùåíèÿ ïëîñêîñòè ïîëÿðèçîâàííîãî ñâåòà. Èçìåíåíèå 18 îñòàòêîâ Ðèñ. 1.17. Ñòðóêòóðà è ïàðàìåòðû à-ñïèðàëè. ïîñëåäíåãî íàõîäèòñÿ â ïðÿìîé çàâèñèìîñòè îò ñòåïåíè ñïèðàëèçàöèè áåëêîâîé ìîëåêóëû. Íå âñå ãëîáóëÿðíûå áåëêè ñïèðàëèçîâàíû íà âñåì ïðîòÿæåíèè ïîëèïåïòèäíîé öåïè. Â ìîëåêóëå áåëêà à-ñïèðàëüíûå ó÷àñòêè ÷åðåäóþòñÿ ñ ëèíåéíûìè. Â ÷àñòíîñòè, åñëè à- è â-öåïè ãåìîãëîáèíà ñïèðàëèçîâàíû, íàïðèìåð, íà 75%, òî ëèçîöèìà—íà 42%, à ïåïñèíà—âñåãî íà 30%. Òàêèì îáðàçîì, ñòàáèëüíîñòü âòîðè÷íîé ñòðóêòóðû îáåñïå÷èâàåòñÿ â îñíîâíîì âîäîðîäíûìè ñâÿçÿìè (îïðåäåëåííûé âêëàä âíîñÿò è ãëàâíîâàëåíòíûå ñâÿçè—ïåïòèäíûå è äèñóëüôèäíûå). Âîäîðîäíàÿ ñâÿçü ïðåäñòàâëÿåò ñîáîé ñëàáîå ýëåêòðîñòàòè÷åñêîå ïðèòÿæåíèå (âçàèìîäåéñòâèå, ñâÿçü) ìåæäó îäíèì ýëåêòðîîòðèöàòåëüíûì àòîìîì (íàïðèìåð, êèñëîðîäîì èëè àçîòîì) è âîäîðîäíûì àòîìîì, êîâàëåíòíî ñâÿçàííûì ñî âòîðûì ýëåêòðîîòðèöàòåëüíûì àòîìîì. Òèïû âîäîðîäíûõ ñâÿçåé ïðåäñòàâëåíû äàëåå. Ïî ñîâðåìåííûì ïðåäñòàâëåíèÿì, âîäîðîäíàÿ ñâÿçü âêëþ÷àåò íå òîëüêî ýëåêòðîñòàòè÷åñêèå ñèëû ïðèòÿæåíèÿ ìåæäó ïîëÿðíûìè ãðóïïàìè (âçàèìî-äåéñòâèå àòîìîâ âîäîðîäà ñ ýëåêòðîîòðèöàòåëüíûìè ýëåìåíòàìè: êèñëîðîäîì, àçîòîì, õëîðîì), íî è ýëåêòðîííûå ñâÿçè òàêîãî æå òèïà, êàê â ðÿäå êîìïëåêñíûõ ñîåäèíåíèé. Âîäîðîäíûå ñâÿçè, ÿâëÿÿñü íåêîâàëåíòíûìè, îòëè÷àþòñÿ ìàëîé ïðî÷íîñòüþ. Òàê, åñëè äëÿ ðàçðûâà õèìè÷åñêèõ ìåæàòîìíûõ ñâÿçåé íåîáõîäèìî çàòðàòèòü îò 84 äî 8400 êÄæ, òî äëÿ ðàçðûâà îäíîé âîäîðîäíîé ñâÿçè òðåáóåòñÿ çàòðàòèòü âñåãî ëèøü 6,3 êÄæ íà 1 ìîëü. Ïîñêîëüêó â áåëêîâîé ìîëåêóëå ÷èñëî âîäîðîäíûõ ñâÿçåé î÷åíü âåëèêî (â îáðàçîâàíèå âîäîðîäíûõ ñâÿçåé âîâëå÷åíû âñå ïåïòèäíûå ãðóïïû), îíè â ñóììå îáåñïå÷èâàþò ñêðó÷èâàíèå ïîëèïåïòèäíîé öåïè â ñïèðàëüíóþ ñòðóêòóðó, ñîîáùàÿ åé êîìïàêòíîñòü è ñòàáèëüíîñòü. Ìåõàíèçì âîçíèêíîâåíèÿ âîäîðîäíûõ ñâÿçåé â ýëåìåíòàðíîé ôîðìå ìîæåò áûòü ïðåäñòàâëåí íà ïðèìåðå âçàèìîäåéñòâèÿ äâóõ ìîëåêóë âîäû (äèïîëè). Â äèïîëå âîäû, êàê èçâåñòíî, èçáûòîê ïîëîæèòåëüíûõ çàðÿäîâ ïðèõîäèòñÿ íà àòîìû âîäîðîäà, à èçáûòîê îòðèöàòåëüíûõ—íà àòîìû êèñëîðîäà. Áëàãîäàðÿ îñîáåííîñòÿì ñòðîåíèÿ àòîìà âîäîðîäà ïðè äîñòàòî÷íîì ñáëèæåíèè äâóõ ìîëåêóë âîäû âîçíèêàåò ýëåêòðîñòàòè÷åñêîå âçàèìîäåéñòâèå ìåæäó àòîìîì êèñëîðîäà îäíîé ìîëåêóëû è àòîìîì âîäîðîäà âòîðîé ìîëåêóëû âîäû. Ñëåäñòâèåì ýòîãî ÿâëÿåòñÿ îñëàáëåíèå ñâÿçè ìåæäó àòîìà-ìè âîäîðîäà è êèñëîðîäà â êàæäîé ìîëåêóëå âîäû è ñîîòâåòñòâåííî âîçíèêíîâåíèå íîâîé, íåïðî÷íîé ñâÿçè (îòìå÷åíà ïóíêòèðîì) ìåæäó àòîìîì âîäîðîäà ïåðâîé ìîëåêóëû è àòîìîì êèñëîðîäà âòîðîé ìîëåêóëû âîäû. Ýòó íåïðî÷íóþ ñâÿçü ïðèíÿòî îáîçíà÷àòü âîäîðîäíîé ñâÿçüþ. Â áåëêîâîé ìîëåêóëå íàèáîëåå âàæíûå âîäîðîäíûå ñâÿçè îáðàçóþòñÿ ìåæäó êîâàëåíòíî ñâÿçàííûì àòîìîì âîäîðîäà, íåñóùèì ÷àñòè÷íûé ïîëîæèòåëüíûé çàðÿä, è îòðèöàòåëüíî çàðÿæåííûì êîâàëåíòíî ñâÿçàííûì àòîìîì êèñëîðîäà. Íèæå ïðåäñòàâëåíû ïðèìåðû âîäîðîäíûõ ñâÿçåé â áåëêîâîé ìîëåêóëå: à) ìåæäó ïåïòèäíûìè öåïÿìè; á) ìåæäó äâóìÿ ãèäðîêñèëüíûìè ãðóïïàìè; â) ìåæäó èîíèçèðîâàííîé ÑîÎÍ-ãðóïïîé è ÎÍ-ãðóï- ïîé òèðîçèíà; ã) ìåæäó ÎÍ-ãðóïïîé ñåðèíà è ïåïòèäíîé ñâÿçüþ. ÍÑ-Ð Ð-ÑÍ / N 0=ñ÷ /ÌÍ ^ì-í-î=ñ Ð-ÑÍ ÕÑÍ-Ð \ ÑÍ-Ð ÑÍ2-0-Í—î=ñ^ â Â çàâèñèìîñòè îò õèìè÷åñêîé ïðèðîäû àòîìà-àêöåïòîðà âîäîðîäíûå ñâÿçè îòëè÷àþòñÿ äðóã îò äðóãà ñòåïåíüþ ïðî÷íîñòè. Î êîëè÷åñòâå âîäîðîäíûõ ñâÿçåé â áåëêîâîé ìîëåêóëå ñóäÿò ïî äàííûì èçîòîïíîãî ìåòîäà, â ÷àñòíîñòè ïî âðåìåíè îáìåíà àòîìîâ âîäîðîäà, ó÷àñòâóþùèõ â îáðàçîâàíèè âîäîðîäíîé ñâÿçè, íà äåéòåðèé (ïðè îáðàáîòêå áåëêà òÿæåëîé âîäîé Á20, â êîòîðîé âìåñòî îáû÷íîãî âîäîðîäà ñîäåðæèòñÿ åãî òÿæåëûé èçîòîï äåéòåðèé). Äðóãîé òèï êîíôèãóðàöèè ïîëèïåïòèäíûõ öåïåé, îáíàðóæåííûé â áåëêàõ âîëîñ, øåëêà, ìûøö è â äðóãèõ ôèáðèëëÿðíûõ áåëêàõ, ïîëó÷èë íàçâàíèå Ð-ñòðóêòóðû. Â ýòîì ñëó÷àå äâå èëè áîëåå ëèíåéíûå ïîëèïåï- òèäíûå öåïè, ðàñïîëîæåííûå ïàðàëëåëüíî èëè, ÷àùå, àíòèïàðàëëåëüíî, ïðî÷íî ñâÿçûâàþòñÿ ìåæöåïî÷å÷íûìè âîäîðîäíûìè ñâÿçÿìè ìåæäó ÊÍ- è ÑÎ-ãðóïïàìè ñîñåäíèõ öåïåé, îáðàçóÿ ñòðóêòóðó òèïà ñêëàä÷àòîãî ñëîÿ (ðèñ. 1.18). Ðèñ. 1.18. â-Ñòðóêòóðà ïîëèïåï òèäíûõ öåïåé. Â ïðèðîäå ñóùåñòâóþò áåëêè, ñòðîåíèå êîòîðûõ, îäíàêî, íå ñîîòâåòñòâóåò íè â-, íè à-ñòðóêòóðå. Òèïè÷íûì ïðèìåðîì òàêèõ áåëêîâ ÿâëÿåòñÿ êîëëàãåí—ôèáðèëëÿðíûé áåëîê, ñîñòàâëÿþùèé îñíîâíóþ ìàññó ñîåäèíèòåëüíîé òêàíè â îðãàíèçìå ÷åëîâåêà è æèâîòíûõ (ñì. ãëàâó 21). Ìåòîäàìè ðåíòãåíîñòðóêòóðíîãî àíàëèçà â íàñòîÿùåå âðåìÿ äîêàçàíî ñóùåñòâîâàíèå åùå äâóõ óðîâíåé ñòðóêòóðíîé îðãàíèçàöèè áåëêîâîé ìîëåêóëû, îêàçàâøèõñÿ ïðîìåæóòî÷íûìè ìåæäó âòîðè÷íîé è òðåòè÷íîé ñòðóêòóðàìè. Ýòî òàê íàçûâàåìûå íàäâòîðè÷íûå ñòðóêòóðû è ñòðóêòóðíûå äîìåíû. Íàäâòîðè÷íûå ñòðóêòóðû ïðåäñòàâëÿþò ñîáîé àãðåãàòû ïîëèïåïòèäíûõ öåïåé, îáëàäàþùèõ ñîáñòâåííîé âòîðè÷íîé ñòðóêòóðîé è îáðàçóþùèõñÿ â íåêîòîðûõ áåëêàõ â ðåçóëüòàòå èõ òåðìîäèíàìè÷åñêîé èëè êèíåòè÷åñêîé ñòàáèëüíîñòè. Òàê, â ãëîáóëÿðíûõ áåëêàõ îòêðûòû (âõâ)-ýëåìåíòû (ïðåäñòàâëåíû äâóìÿ ïàðàëëåëüíûìè â-öåïÿìè, ñâÿçàííûìè ñåãìåíòîì õ), âàâàâ-ýëåìåíòû (ïðåäñòàâëåíû äâóìÿ ñåãìåíòàìè à-ñïèðàëè, âñòàâëåííûìè ìåæäó òðåìÿ ïàðàëëåëüíûìè â-öåïÿìè) è äð. Â áîëüøèõ ãëîáóëÿðíûõ áåëêàõ èíîãäà ñîäåðæàòñÿ íåîäèíàêîâûå ñòðóêòóðíûå äîìåíû, âûïîëíÿþùèå ðàçíûå ôóíêöèè, êàê è îäíîòèïíûå äîìåíû â ïðåäåëàõ îäíîãî ìîíîìåðíîãî áåëêà, îáðàçóþùèåñÿ, âåðîÿòíåå âñåãî, êàê ðåçóëüòàò âëèÿíèÿ ãåíîâ â ïåðâîì ñëó÷àå èëè äóïëèêàöèè ãåíîâ — âî âòîðîì. Äîìåíû ñîçäàþòñÿ îáúåäèíåíèåì è ÷åðåäîâàíèåì à-ñïèðàëåé è â-ñëîåâ, ìåæäó êîòîðûìè îòêðûâàþòñÿ áîëåå ðûõëûå ñòðóêòóðû (ðèñ. 1.19). Äîìåí—ýòî êîìïàêòíàÿ ãëîáóëÿðíàÿ ñòðóêòóðíàÿ åäèíèöà âíóòðè ïîëèïåïòèäíîé öåïè. Äîìåíû ìîãóò âûïîëíÿòü ðàçíûå ôóíêöèè è ïîäâåðãàòüñÿ ñêëàäûâàíèþ (ñâåðòûâàíèþ) â íåçàâèñèìûå êîìïàêòíûå ãëîáóëÿðíûå ñòðóêòóðíûå åäèíèöû, ñîåäèíåííûå ìåæäó ñîáîé ãèáêèìè ó÷àñòêàìè âíóòðè áåëêîâîé ìîëåêóëû. Îòêðûòî ìíîãî áåëêîâ (íàïðèìåð, èììóíîãëîáóëèíû), ñîñòîÿùèõ èç ðàçíûõ ïî ñòðóêòóðå è ôóíêöèÿì äîìåíîâ, êîäèðóåìûõ ðàçíûìè ãåíàìè. Òðåòè÷íàÿ ñòðóêòóðà áåëêà Ïîä òðåòè÷íîé ñòðóêòóðîé áåëêà ïîäðàçóìåâàþò ïðîñòðàíñòâåííóþ îðèåíòàöèþ ïîëèïåïòèäíîé ñïèðàëè èëè ñïîñîá óêëàäêè ïîëèïåïòèäíîé öåïè â îïðåäåëåííîì îáúåìå. Ïîñêîëüêó íè ïåðâè÷íàÿ ñòðóêòóðà, íè òèïû ñïèðàëåé èëè ñî÷åòàíèÿ ñïèðàëüíûõ è ëèíåéíûõ ó÷àñòêîâ ïîëèïåïòèäíîé Ðèñ. 1.19. Äîìåííîå ñòðîåíèå ãëîáóëÿðíûõ áåëêîâ (ïî À. À. Áîëäûðåâó). à - â-ñóáúåäèíèöà ãåìîãëîáèíà; á - êîíñòàíòíûé äîìåí èììóíîãëîáóëèíà; â - ôëàâîäîêñèí; ã - ëèçîöèì êóðèíîãî ÿéöà. öåïè íå äàþò ïðåäñòàâëåíèÿ îá îáúåìå, ôîðìå ïîëèïåïòèäíîé öåïè, ïåðåä èññëåäîâàòåëåì âñåãäà ñòîèò íåîáõîäèìîñòü îïðåäåëåíèÿ òðåõìåðíîé èëè ïðîñòðàíñòâåííîé êîíôèãóðàöèè áåëêà. Îñíîâíóþ ðîëü â ðåøåíèè ýòèõ çàäà÷ ñûãðàë ðåíòãåíîñòðóêòóðíûé àíàëèç ñ âûñîêîé ðàçðåøàþùåé ñïîñîáíîñòüþ. Êàê áûëî îòìå÷åíî, ìåòîä óñïåøíî ðåøàåò äâå ãëàâíûå ïðîáëåìû õèìèè áåëêîâ: çàêîíîìåðíîñòü ïîñëåäîâàòåëüíîñòåé àìèíîêèñëîòíûõ îñòàòêîâ â ïîëèïåïòèäå è çàêîíîìåðíîñòü êîíôèãóðàöèè ìîëåêóëû áåëêà. Ìåæàòîìíûå ðàññòîÿíèÿ â ìîëåêóëàõ îðãàíè÷åñêèõ âåùåñòâ ñîñòàâëÿþò 0,1—0,2 íì, à ìàêñèìàëüíàÿ ðàçðåøàþùàÿ ñïîñîáíîñòü ñîâðåìåííûõ àïïàðàòîâ ðàâíà 0,2 íì. Ýòî íå ïîçâîëÿåò óñòàíîâèòü ìåñòîïîëîæåíèå êàæäîãî àòîìà, õîòÿ âïîëíå ìîãóò áûòü ðàçëè÷èìû îòäåëüíûå ñî÷åòàíèÿ àòîìîâ, îñîáåííî ïðè ââåäåíèè â ìîëåêóëó áåëêîâ àòîìîâ òÿæåëûõ ìåòàëëîâ Ðèñ. 1.20. Ìîäåëü òðåòè÷íîé ñòðóêòóðû ìîëåêóëû ìèîãëîáèíà (ïî Äæ. Êåíäðüþ). Ëàòèíñêèìè áóêâàìè îáîçíà÷åíû ñòðóêòóðíûå äîìåíû, êðàñíûì öâåòîì — ãåì. (ïîñëåäíèå áëàãîäàðÿ ñâîåé âûñîêîé ýëåêòðîííîé ïëîòíîñòè èñïîëüçóþòñÿ â êà÷åñòâå òî÷åê îòñ÷åòà ïðè ìàòåìàòè÷åñêîé îáðàáîòêå ðåíòãåíîãðàìì). Ïåðâûì áåëêîì, òðåòè÷íàÿ ñòðóêòóðà êîòîðîãî áûëà âûÿñíåíà Äæ. Êåíäðüþ íà îñíîâàíèè ðåíòãåíîñòðóêòóðíîãî àíàëèçà, îêàçàëñÿ ìèî- ãëîáèí êàøàëîòà. Ýòî ñðàâíèòåëüíî íåáîëüøîé áåëîê ñ ìîë. ì. 16700, ñîäåðæàùèé 153 àìèíîêèñëîòíûõ îñòàòêà (ïîëíîñòüþ âûÿñíåíà ïåðâè÷íàÿ ñòðóêòóðà), ïðåäñòàâëåííûé îäíîé ïîëèïåïòèäíîé öåïüþ. Îñíîâíàÿ ôóíêöèÿ ìèîãëîáèíà — ïåðåíîñ êèñëîðîäà â ìûøöàõ. Ïîëèïåïòèäíàÿ öåïü ìèîãëîáèíà (ðèñ. 1.20) ïðåäñòàâëåíà â âèäå èçîãíóòîé òðóáêè, êîìïàêòíî óëîæåííîé âîêðóã ãåìà (íåáåëêîâûé êîìïîíåíò, ñîäåðæàùèé æåëåçî; ñì. ãëàâó 2). Íà ïðîòÿæåíèè ïîñëåäíèõ äåñÿòèëåòèé â ñâÿçè ñ ïîâûøåíèåì ðàçðåøàþùåé ñïîñîáíîñòè ðåíòãåíîñòðóêòóðíîãî ìåòîäà áûëà ðàñøèôðîâàíà òðåòè÷íàÿ ñòðóêòóðà áîëåå 1000 áåëêîâ, â òîì ÷èñëå ãåìîãëîáèíà, ïåïñèíà, õèìîòðèïñèíà, ðèáîíóêëåàçû, ëèçîöèìà, òðèïñèíà è åãî èíãèáèòîðà, ðÿäà ôðàãìåíòîâ èììóíîãëîáóëèíîâ ÷åëîâåêà, öèòîõðîìà Ñ, êàðáîàíãèäðàçû ÷åëîâåêà, àñïàðòàòàìèíîòðàíñôåðàçû, èíñóëèíà è äð. Ïðèìåðû òðåõìåðíîé ñòðóêòóðû íåêîòîðûõ èç íèõ ïðåäñòàâëåíû íà ðèñ. 1.21. Ðåíòãåíîñòðóêòóðíûé àíàëèç ïîçâîëÿåò îïðåäåëèòü êîíôîðìàöèþ è õîä ïîëèïåïòèäíîé öåïè â ïðîñòðàíñòâå, ïîýòîìó äëÿ êàæäîãî áåëêà ìîæåò áûòü ïîñòðîåíà îáúåìíàÿ ìîäåëü, îòðàæàþùàÿ ìåñòîïîëîæåíèå ëèíåéíûõ è ñïèðàëèçîâàííûõ ó÷àñòêîâ. Ïðè èçó÷åíèè ãëîáóëÿðíûõ áåëêîâ áûëî ïîêàçàíî, ÷òî ïðîñòðàíñòâåííàÿ ñòðóêòóðà áåëêîâ â ñèëüíîé ñòåïåíè çàâèñèò îò ðÿäà ôàêòîðîâ, â ÷àñòíîñòè îò èîííîé ñèëû è ðÍ ðàñòâîðà, òåìïåðàòóðû è ò.ä. Íîâåéøèå ìåòîäû äèôðàêöèè ðåíòãåíîâñêèõ ëó÷åé ïîçâîëèëè ðàñøèôðîâàòü êðèñòàëëè÷åñêóþ ñòðóêòóðó áîëåå 100 ôåðìåíòîâ. Äëÿ âûÿñíåíèÿ òðåõìåðíîé ñòðóêòóðû áåëêîâ â ïîñëåäíåå âðåìÿ óñïåøíî ïðèìåíÿþòñÿ òàêæå ìåòîäû íèçêîòåìïåðàòóðíîé âû÷èñëèòåëüíîé òåõíèêè, à òàêæå ìàòåìàòè÷åñêèå è êîìïüþòåðíûå ìåòîäû îïðåäåëåíèÿ îáúåìíîé ñòðóêòóðû íà îñíîâàíèè äàííûõ ïîñëåäîâàòåëüíîñòåé àìèíîêèñëîò. Â íàñòîÿùåå âðåìÿ ïîëó÷åíû áåññïîðíûå äîêàçàòåëüñòâà, ÷òî â ñòàáèëèçàöèè ïðîñòðàíñòâåííîé ñòðóêòóðû áåëêîâ, ïîìèìî êîâàëåíòíûõ ñâÿçåé (ïåïòèäíûå è äèñóëüôèäíûå ñâÿçè), îñíîâíóþ ðîëü èãðàþò òàê íàçûâàåìûå íåêîâàëåíòíûå ñâÿçè (ðèñ. 1.22). Ê ýòèì ñâÿçÿì îòíîñÿòñÿ âîäîðîäíûå ñâÿçè, ýëåêòðîñòàòè÷åñêèå âçàèìîäåéñòâèÿ çàðÿæåííûõ ãðóïï, ìåæìîëåêó- ëÿðíûå âàí-äåð-âààëüñîâû ñèëû, âçàèìîäåéñòâèÿ íåïîëÿðíûõ áîêîâûõ ðàäèêàëîâ àìèíîêèñëîò, òàê íàçûâàåìûå ãèäðîôîáíûå âçàèìîäåéñòâèÿ è ò.ä. Ïî ñîâðåìåííûì ïðåäñòàâëåíèÿì, òðåòè÷íàÿ ñòðóêòóðà áåëêà ïîñëå çàâåðøåíèÿ åãî ñèíòåçà â ðèáîñîìàõ (ñì. ãëàâó 14) ôîðìèðóåòñÿ ñîâåðøåííî àâòîìàòè÷åñêè, ñàìîïðîèçâîëüíî è ïîëíîñòüþ ïðåäîïðåäåëÿåòñÿ ïåðâè÷íîé ñòðóêòóðîé. Îñíîâíîé äâèæóùåé ñèëîé â âîçíèêíîâåíèè òðåõìåðíîé ñòðóêòóðû ÿâëÿåòñÿ âçàèìîäåéñòâèå ðàäèêàëîâ àìèíîêèñëîò ñ ìîëåêóëàìè âîäû. Ïðè ýòîì íåïîëÿðíûå ãèäðîôîáíûå ðàäèêàëû àìèíîêèñëîò êàê áû ïîãðóæàþòñÿ âíóòðü áåëêîâîé ìîëåêóëû, îáðàçóÿ òàì ñóõèå çîíû, â òî âðåìÿ êàê ïîëÿðíûå ðàäèêàëû îêàçûâàþòñÿ îðèåíòèðîâàííûìè â ñòîðîíó âîäû. Â êàêîé-òî ìîìåíò âîçíèêàåò òåðìîäèíàìè÷åñêè íàèáîëåå âûãîäíàÿ ñòàáèëüíàÿ êîíôîðìàöèÿ ìîëåêóëû. Â òàêîé ôîðìå áåëêîâàÿ ìîëåêóëà õàðàêòåðèçóåòñÿ ìèíèìàëüíîé ñâîáîäíîé ýíåðãèåé. Ìîëåêóëû áåëêîâ â âîäíûõ ðàñòâîðàõ îáû÷íî ïðèíèìàþò ðÿä ñòàáèëüíûõ êîíôîðìàöèé, èíäóöèðóåìûõ íå òîëüêî èçìåíåíèÿìè ðÍ è òåìïåðàòóðû, íî è íèçêîìîëåêóëÿðíûìè ñîåäèíåíèÿìè. Ðàçëè÷àþò äâå îñíîâíûå ôîðìû êîíôîðìàöèé: Ò-ôîðìó (îò àíãë. 1åïçå¸—íàïðÿæåííàÿ) è Ê-ôîðìó (îò àíãë. ãå1àõå¸—ðàñ- Ðèñ. 1.22. Òèïû íåêîâàëåíòíûõ ñâÿçåé, ñòàáèëèçèðóþùèõ òðåòè÷íóþ ñòðóêòóðó áåëêà. à - ýëåêòðîñòàòè÷åñêîå âçàèìîäåéñòâèå; á - âîäîðîäíàÿ ñâÿçü; â - ãèäðîôîáíûå âçàèìîäåéñòâèÿ íåïîëÿðíûõ ãðóïï; ã - äèïîëü-äèïîëüíûå âçàèìîäåéñòâèÿ; ä - äèñóëüôèäíàÿ (êîâàëåíòíàÿ) ñâÿçü. ñëàáëåííàÿ). Ìåæäó ýòèìè ôîðìàìè îñóùåñòâëÿþòñÿ ïåðåõîäû, ñîîòâåòñòâåííî îòðàæàþùèåñÿ â áèîëîãè÷åñêèõ ñâîéñòâàõ. Â ïðîöåññå óêëàäêè ñèíòåçèðîâàííîé ïîëèïåïòèäíîé öåïè, ïîëó÷èâøåì íàçâàíèå ôîëäèíãà—ôîðìèðîâàíèå íàòèâíîé ïðîñòðàíñòâåííîé ñòðóêòóðû, â êëåòêàõ ïðîèñõîäèò îòáîð èç ìíîæåñòâà ñòåðè÷åñêè âîçìîæíûõ ñîñòîÿíèé îäíîé-åäèíñòâåííîé ñòàáèëüíîé è áèîëîãè÷åñêè àêòèâíîé êîí-ôîðìàöèè, îïðåäåëÿåìîé, âåðîÿòíåå âñåãî, ïåðâè÷íîé ñòðóêòóðîé. Îïèñàí ðÿä íàñëåäñòâåííûõ çàáîëåâàíèé ÷åëîâåêà, ðàçâèòèå êîòîðûõ ñâÿçûâàþò ñ íàðóøåíèåì âñëåäñòâèå ìóòàöèé ïðîöåññà ôîëäèíãà (ïèãìåíòîçû, ôèáðîçû è äð.). Ïîýòîìó â íàñòîÿùåå âðåìÿ ïðèñòàëüíîå âíèìàíèå èññëåäîâàòåëåé ïðèêîâàíî ê âûÿñíåíèþ çàâèñèìîñòè ìåæäó àìèíîêèñëîòíîé ïîñëåäîâàòåëüíîñòüþ ñèíòåçèðîâàííîé â êëåòêå ïîëèïåïòèäíîé öåïè (ïåðâè÷íàÿ ñòðóêòóðà) è ôîðìèðîâàíèåì ïðîñòðàíñòâåííîé òðåõìåðíîé ñòðóêòóðû, îáåñïå÷èâàþùåé áåëêîâîé ìîëåêóëå åå íàòèâíûå ñâîéñòâà. Èìååòñÿ íåìàëî ýêñïåðèìåíòàëüíûõ äîêàçàòåëüñòâ, ÷òî ýòîò ïðîöåññ íå ÿâëÿåòñÿ àâòîìàòè÷åñêèì, êàê ïðåäïîëàãàëîñü ðàíåå, è, âåðîÿòíåå âñåãî, ðåãóëèðóåòñÿ è êîíòðîëèðóåòñÿ òàêæå âíóòðèêëåòî÷íûìè ìîëåêóëÿðíûìè ìåõàíèçìàìè, äåòàëè êîòîðûõ ïîêà ïîëíîñòüþ íå ðàñêðûòû. Èç êëåòîê âûäåëåíî íåñêîëüêî êëàññîâ áåëêîâ, íàçâàííûõ øàïåðîíàìè, èëè áåëêàìè òåïëîâîãî øîêà, êîòîðûå ðàñïîëàãàþòñÿ ìåæäó Ê-êîíöåâûì ñèãíàëüíûì ïåïòèäîì è ìàòðè÷íûì áåëêîì. Ïðåäïîëàãàåòñÿ, ÷òî îñíîâíûìè ôóíêöèÿìè øàïåðîíîâ ÿâëÿþòñÿ ñïîñîáíîñòü ïðåäîòâðàùàòü îáðàçîâàíèå èç ïîëèïåïòèäíîé öåïè íåñïåöèôè÷åñêèõ (õàîòè÷íûõ) áåñïîðÿäî÷íûõ êëóáêîâ, èëè àãðåãàòîâ áåëêîâ, è îáåñïå÷åíèå äîñòàâêè (òðàíñïîðòà) èõ ê ñóáêëåòî÷íûì ìèøåíÿì, ñîçäàâàÿ óñëîâèÿ äëÿ çàâåðøåíèÿ ñâåðòûâàíèÿ áåëêîâîé ìîëåêóëû. Ýòè ðåçóëüòàòû íàâîäÿò íà ìûñëü î âîçìîæíîñòè ñóùåñòâîâàíèÿ «âòîðîé ïîëîâèíû ãåíåòè÷åñêîãî êîäà», îïðåäåëÿÿ òåì ñàìûì ïîâûøåííûé èíòåðåñ èññëåäîâàòåëåé ê ïðîáëåìå ñâåðòûâàíèÿ ïîëèïåïòèäíîé öåïè è ôîðìèðîâàíèÿ åå íàòèâíîé ïðîñòðàíñòâåííîé êîíôîðìàöèè. Òàêèì îáðàçîì, ëèíåéíàÿ îäíîìåðíàÿ ñòðóêòóðà ïîëèïåïòèäíîé öåïè (ò.å. ïîñëåäîâàòåëüíîñòü àìèíîêèñëîòíûõ îñòàòêîâ, îáóñëîâëåííàÿ êîäîì áåëêîâîãî ñèíòåçà) íàäåëåíà èíôîðìàöèåé äðóãîãî òèïà—êîíôîðìàöèîí- íîé, êîòîðàÿ ïðåäñòàâëÿåò ñîáîé îáðàçîâàíèå áåëêîâîé ìîëåêóëû ñòðîãî çàäàííîé ôîðìû ñ îïðåäåëåííûì ïðîñòðàíñòâåííûì ðàñïîëîæåíèåì îòäåëüíûõ åå ÷àñòåé. Äðóãèìè ñëîâàìè, òðåòè÷íàÿ—îáúåìíàÿ—ñòðóêòóðà áåë-êîâîé ìîëåêóëû äåòåðìèíèðîâàíà àìèíîêèñëîòíîé ïîñëåäîâàòåëüíîñòüþ ïîëèïåïòèäíîé öåïè, à áîëåå êîíêðåòíî—ðàçìåðîì, ôîðìîé è ïîëÿðíîñòüþ ðàäèêàëîâ àìèíîêèñëîòíûõ îñòàòêîâ. Ýòè ïðåäñòàâëåíèÿ ìîãóò ñëóæèòü îñíîâîé äëÿ ïðåäñêàçàíèÿ êîíôîðìàöèè áåëêîâîé ìîëåêóëû íà îñíîâàíèè àìèíîêèñëîòíîé ïîñëåäîâàòåëüíîñòè. Ñëåäóåò óêàçàòü, îäíàêî, ÷òî äî ñèõ ïîð ïðåäñòàâëÿåòñÿ èíòðèãóþùåé çàãàäêîé ìåõàíèçì ýòîé òåñíîé è òîíêîé ñâÿçè ìåæäó àìèíîêèñëîòíîé ïîñëåäîâàòåëüíîñòüþ è òðåõìåðíîé ñòðóêòóðîé áåëêîâîé ìîëåêóëû. Îêàçûâàåòñÿ, èíîãäà ïîëèïåïòèäû ïî÷òè ñ îäèíàêîâûìè ïîñëåäîâàòåëüíîñòÿìè îáðàçóþò ðàçíûå ñòðóêòóðû è, íàîáîðîò, ïîëèïåïòèäû ñ ðàçíûìè ïîñëåäîâàòåëüíîñòÿìè ôîðìèðóþò îäèíàêîâóþ òðåõìåðíóþ ñòðóêòóðó. Â ñâîþ î÷åðåäü òðåõìåðíàÿ ñòðóêòóðà áåëêîâîé ìîëåêóëû òàêæå ñîäåðæèò èíôîðìàöèþ, íî óæå ñîâåðøåííî íîâîãî òèïà, à èìåííî ôóíêöèîíàëü-íóþ, êîòîðóþ àêàä. Â.À. Ýíãåëüãàðäò íàçâàë èíòðàìîëåêóëÿðíîé èíôîðìàöèåé. Êàê áóäåò ïîêàçàíî äàëåå, âñå áèîëîãè÷åñêèå ñâîéñòâà áåëêîâ (êàòàëèòè÷åñêèå, ãîðìîíàëüíûå, àíòèãåííûå è äð.) ñâÿçàíû ñ ñîõðàííîñòüþ èõ òðåòè÷íîé ñòðóêòóðû, êîòîðóþ ïðèíÿòî íàçûâàòü íàòèâíîé êîíôîðìàöèåé. Ëþáûå âîçäåéñòâèÿ (òåðìè÷åñêèå, ôèçè÷åñêèå, õèìè÷åñêèå), ïðèâîäÿùèå ê íàðóøåíèþ ýòîé êîíôîðìàöèè ìîëåêóëû (ðàçðûâ âîäîðîäíûõ è äðóãèõ íåêîâàëåíòíûõ ñâÿçåé), ñîïðîâîæäàþòñÿ ÷àñòè÷íîé èëè ïîëíîé ïîòåðåé áåëêîì åãî áèîëîãè÷åñêèõ ñâîéñòâ. ×åòâåðòè÷íàÿ ñòðóêòóðà áåëêà Ïîä ÷åòâåðòè÷íîé ñòðóêòóðîé ïîäðàçóìåâàþò ñïîñîá óêëàäêè â ïðîñòðàíñòâå îòäåëüíûõ ïîëèïåïòèäíûõ öåïåé, îáëàäàþùèõ îäèíàêîâîé (èëè ðàçíîé) ïåðâè÷íîé, âòîðè÷íîé èëè òðåòè÷íîé ñòðóêòóðîé, è ôîðìèðîâàíèå åäèíîãî â ñòðóêòóðíîì è ôóíêöèîíàëüíîì îòíîøåíèÿõ ìàêðîìîëåêóëÿðíî- ãî îáðàçîâàíèÿ. Ìíîãèå ôóíêöèîíàëüíûå áåëêè ñîñòîÿò èç íåñêîëüêèõ ïîëèïåïòèäíûõ öåïåé, ñîåäèíåííûõ íå ãëàâíîâàëåíòíûìè ñâÿçÿìè, à íåêîâàëåíòíûìè (àíàëîãè÷íûìè òåì, êîòîðûå îáåñïå÷èâàþò ñòàáèëüíîñòü òðåòè÷íîé ñòðóêòóðû). Êàæäàÿ îòäåëüíî âçÿòàÿ ïîëèïåïòèäíàÿ öåïü, ïîëó÷èâøàÿ íàçâàíèå ïðîòîìåðà, ìîíîìåðà èëè ñóáúåäèíèöû, ÷àùå âñåãî íå îáëàäàåò áèîëîãè÷åñêîé àêòèâíîñòüþ. Ýòó ñïîñîáíîñòü áåëîê ïðèîáðåòàåò ïðè îïðåäåëåííîì ñïîñîáå ïðîñòðàíñòâåííîãî îáúåäèíåíèÿ âõîäÿùèõ â åãî ñîñòàâ ïðîòîìåðîâ, ò.å. âîçíèêàåò íîâîå êà÷åñòâî, íå ñâîéñòâåííîå ìîíîìåðíîìó áåëêó. Îáðàçîâàâøóþñÿ ìîëåêóëó ïðèíÿòî íàçûâàòü îëèãîìåðîì (èëè ìóëüòèìåðîì). Îëèãîìåðíûå áåëêè ÷àùå ïîñòðîåíû èç ÷åòíîãî ÷èñëà ïðîòîìåðîâ (îò 2 äî 4, ðåæå îò 6 äî 8) ñ îäèíàêîâûìè èëè ðàçíûìè ìîëåêóëÿðíûìè ìàññàìè — îò íåñêîëüêèõ òûñÿ÷ äî ñîòåí òûñÿ÷. Â ÷àñòíîñòè, ìîëåêóëà ãåìîãëîáèíà ñîñòîèò èç äâóõ îäèíàêîâûõ à- è äâóõ â-ïîëèïåïòèäíûõ öåïåé, ò.å. ïðåäñòàâëÿåò ñîáîé òåòðàìåð. Íà ðèñ. 1.23 ïðåäñòàâëåíà ñòðóêòóðà ìîëåêóëû ãåìîãëîáèíà, à íà ðèñ. 1.24 õîðîøî âèäíî, ÷òî ìîëåêóëà ãåìîãëîáèíà ñîäåðæèò ÷åòûðå ïîëèïåïòèäíûå öåïè, êàæäàÿ èç êîòîðûõ îêðóæàåò ãðóïïó ãåìà — ïèãìåíòà, ïðèäàþùåãî êðîâè åå õàðàêòåðíûé êðàñíûé öâåò (ñì. ãëàâó 2). Â îïðåäåëåííûõ óñëîâèÿõ (ïðèñóòñòâèå ñîëåé, 8Ì ìî÷åâèíû èëè ðåçêèå èçìåíåíèÿ ðÍ) ìîëåêóëà ãåìîãëîáèíà îáðàòèìî äèññîöèèðóåò íà äâå àè äâå â-öåïè. Ýòà äèññîöèàöèÿ îáóñëîâëåíà ðàçðûâîì âîäîðîäíûõ ñâÿçåé. Ïîñëå óäàëåíèÿ ñîëåé èëè ìî÷åâèíû ïðîèñõîäèò àâòîìàòè÷åñêàÿ àññîöèàöèÿ èñõîäíîé ìîëåêóëû ãåìîãëîáèíà (ðèñ. 1.25). Êëàññè÷åñêèì ïðèìåðîì îëèãîìåðíîé ìîëåêóëû, èëè íàäìîëåêóëÿðíîé ñòðóêòóðû, ÿâëÿåòñÿ âèðóñ òàáà÷íîé ìîçàèêè, ïðåäñòàâëÿþùèé ñîáîé ãèãàíòñêóþ ìîëåêóëó ñ ìîë. ì. îêîëî 40 • 106. Îí ñîñòîèò èç îäíîé ìîëåêóëû ÐÍÊ (ñì. ãëàâó 3) è 2130 áåëêîâûõ ñóáúåäèíèö, ìàññà êàæäîé èç êîòîðûõ ñîñòàâëÿåò 17500. Äëèíà âèðóñà ïðèìåðíî 300 íì, øèðèíà—îêîëî 17 íì. ÐÍÊ âèðóñà èìååò ñïèðàëåîáðàçíóþ ôîðìó. Âîêðóã ÐÍÊ íàíèçàíû áåëêîâûå ÷àñòèöû, îáðàçóþùèå ãèãàíòñêóþ íàäìîëåêóëÿðíóþ ñïèðàëüíóþ ñòðóêòóðó, â êîòîðîé íàñ÷èòûâàåòñÿ îêîëî 130 âèòêîâ (ðèñ. 1.26). Óäèâèòåëüíîé îñîáåííîñòüþ âèðóñà ÿâëÿåòñÿ òî, ÷òî ïîñëå ðàçúåäèíåíèÿ ñîîòâåòñòâóþùèìè ïðèåìàìè (äîáàâëåíèå äåòåðãåíòà) ÐÍÊ è áåëêîâûõ ñóáúåäèíèö è ïîñëåäóþùåãî èõ ñìåøèâàíèÿ (ñ ïðåäâàðèòåëüíûì óäàëåíèåì äåòåðãåíòà) íàáëþäàþòñÿ ïîëíàÿ ðåãåíåðàöèÿ ÷åòâåðòè÷íîé ñòðóêòóðû, âîññòàíîâëåíèå âñåõ ôèçè÷åñêèõ ïàðàìåòðîâ è áèîëîãè÷åñêèõ ôóíêöèé (èíôåêòèâ- íàÿ ñïîñîáíîñòü âèðóñà). Ïîäîáíàÿ òî÷íîñòü ïðîöåññà ñïîíòàííîé ñàìîñáîðêè âèðóñà îáåñïå÷èâàåòñÿ, âåðîÿòíåå âñåãî, èíôîðìàöèåé, ñîäåðæàùåéñÿ â ïåðâè÷íîé ñòðóêòóðå ìîëåêóëû ÐÍÊ è áåëêîâûõ ñóáúåäèíèö. Òàêèì îáðàçîì, ïîñëåäîâàòåëüíîñòü àìèíîêèñëîò ñîäåðæèò â ñåáå èíôîðìàöèþ, êîòîðàÿ ðåàëèçóåòñÿ íà âñåõ óðîâíÿõ ñòðóêòóðíîé îðãàíèçàöèè áåëêîâ. Ìíîãèå ôåðìåíòû òàêæå îáëàäàþò ÷åòâåðòè÷íîé ñòðóêòóðîé, íàïðèìåð ôîñôîðèëàçà à, ñîñòîÿùàÿ èç äâóõ èäåíòè÷íûõ ñóáúåäèíèö, â êàæäîé èç êîòîðûõ ïî äâå ïåïòèäíûå öåïè. Âñÿ ìîëåêóëà ôîñôîðèëàçû à, òàêèì îáðàçîì, ïðåäñòàâëÿåò ñîáîé òåòðàìåð. Îòäåëüíûå ñóáúåäèíèöû ÷àùå âñåãî íå îáëàäàþò êàòàëèòè÷åñêîé àêòèâíîñòüþ; âîîáùå ðåãóëÿòîðíûå ôåðìåíòû äà1 Ðèñ. 1.26. Ñàìîñáîðêà âèðóñà òàáà÷íîé ìîçàèêè. (ñì. ãëàâó 4) èìåþò ÷åòâåðòè÷íóþ îëèãîìåðíóþ ñòðóêòóðó. Îíè íàäåëåíû ôóíêöèåé îáåñïå÷åíèÿ â êëåòêå òðåáóåìûõ ñêîðîñòåé õèìè÷åñêèõ ðåàêöèé. Íàèáîëåå èçó÷åííûì îëèãîìåðíûì ôåðìåíòîì ÿâëÿåòñÿ ëàêòàòäåãèäðî- ãåíàçà (îíà êàòàëèçèðóåò îáðàòèìîå ïðåâðàùåíèå ïèðîâèíîãðàäíîé êèñëîòû â ìîëî÷íóþ), ñîäåðæàùàÿ äâà òèïà ïîëèïåïòèäíûõ öåïåé: Í—ñåðäå÷íûé òèï (îò àíãë. Üåàé—ñåðäöå) è Ì—ìûøå÷íûé òèï (îò àíãë. øèçñ1å—ìûøöà) — è ñîñòîÿùàÿ èç 4 ñóáúåäèíèö. Ýòîò ôåðìåíò áëàãîäàðÿ ðàçëè÷íûì ñî÷åòàíèÿì ñóáúåäèíèö ìîæåò ñóùåñòâîâàòü â 5 ôîðìàõ. Òàêèå ôåðìåíòû ïîëó÷èëè íàçâàíèå èçîôåðìåíòîâ, èëè, â ñîîòâåòñòâèè ñ íîâîé êëàññèôèêàöèåé, ìíîæåñòâåííûõ ôîðì ôåðìåíòîâ (ñì. ãëàâó 4). Ê íàñòîÿùåìó âðåìåíè ñóáúåäèíè÷íàÿ ñòðóêòóðà îáíàðóæåíà ó íåñêîëüêèõ ñîòåí áåëêîâ. Îäíàêî òîëüêî äëÿ íåìíîãèõ áåëêîâ, â òîì ÷èñëå äëÿ ìîëåêóëû ãåìîãëîáèíà, ìåòîäîì ðåíòãåíîñòðóêòóðíîãî àíàëèçà ðàñøèôðîâàíà ÷åòâåðòè÷íàÿ ñòðóêòóðà . Îñíîâíûìè ñèëàìè, ñòàáèëèçèðóþùèìè ÷åòâåðòè÷íóþ ñòðóêòóðó, ÿâëÿþòñÿ íåêîâàëåíòíûå ñâÿçè ìåæäó êîíòàêòíûìè ïëîùàäêàìè ïðîòîìåðîâ, êîòîðûå âçàèìîäåéñòâóþò äðóã ñ äðóãîì ïî òèïó êîìïëåìåíòàðíîñòè—óíèâåðñàëüíîìó ïðèíöèïó, ñâîéñòâåííîìó æèâîé ïðèðîäå. Ñòðóêòóðà áåëêà ïîñëå åãî ñèíòåçà â ðèáîñîìå ìîæåò ÷àñòè÷íî ïîäâåðãàòüñÿ ìîäèôèêàöèè (ïîñòòðàíñëÿöèîííûé ïðîöåññèíã): íàïðèìåð, ïðè ïðåâðàùåíèè ïðåäøåñòâåííèêîâ ðÿäà ôåðìåíòîâ èëè ãîðìîíîâ (èíñóëèí). Òàêèì îáðàçîì, èìåþòñÿ âñå îñíîâàíèÿ äëÿ ïîäòâåðæäåíèÿ ìíåíèÿ î ñóùåñòâîâàíèè 4 óðîâíåé ñòðóêòóðíîé îðãàíèçàöèè áåëêîâ. Áîëåå òîãî, êàæäûé èíäèâèäóàëüíûé áåëîê õàðàêòåðèçóåòñÿ óíèêàëüíîé ñòðóêòóðîé, îáåñïå÷èâàþùåé óíèêàëüíîñòü åãî ôóíêöèé. Ïîýòîìó âûÿñíåíèå ñòðóêòóðû ðàçíîîáðàçíûõ áåëêîâ ìîæåò ñëóæèòü êëþ÷îì ê ïîçíàíèþ ïðèðîäû æèâûõ ñèñòåì è ñîîòâåòñòâåííî ñóùíîñòè æèçíè. Íà ýòîì ïóòè íàó÷íîãî ïîèñêà ìîãóò áûòü ðåøåíû òàêæå ìíîãèå ïðîáëåìû íàñëåäñòâåííûõ çàáîëåâàíèé ÷åëîâåêà, â îñíîâå êîòîðûõ ëåæàò äåôåêòû ñòðóêòóðû è áèîñèíòåçà áåëêîâ. Íåêîòîðûå èññëåäîâàòåëè ñêëîííû ðàññìàòðèâàòü, è íå áåç îñíîâàíèÿ, ñóùåñòâîâàíèå ïÿòîãî óðîâíÿ ñòðóêòóðíîé îðãàíèçàöèè áåëêîâ. Ðå÷ü èäåò î ïîëèôóíêöèîíàëüíûõ ìàêðîìîëåêóëÿðíûõ êîìïëåêñàõ, èëè àññîöèàòàõ èç ðàçíûõ ôåðìåíòîâ, ïîëó÷èâøèõ íàçâàíèå ìåòàáîëè÷åñêèõ îëèãîìåðîâ, èëè ìåòàáîëîíîâ, è êàòàëèçèðóþùèõ âåñü ïóòü ïðåâðàùåíèé ñóáñòðàòà (ñèíòå- òàçû âûñøèõ æèðíûõ êèñëîò, ïèðóâàòäåãèäðîãåíàçíûé êîìïëåêñ, äûõàòåëüíàÿ öåïü). ÊËÀÑÑÈÔÈÊÀÖÈß ÁÅËÊÎÂ Â íàñòîÿùåå âðåìÿ åùå íå ðàçðàáîòàíà ñòðîéíàÿ ñèñòåìà íîìåíêëàòóðû è êëàññèôèêàöèè áåëêîâ. Òðàäèöèîííàÿ êëàññèôèêàöèÿ áåëêîâ ïî ãðóïïàì, îñíîâàííàÿ, ñêîðåå, íà ñëó÷àéíûõ ïîêàçàòåëÿõ (ôèçèêî-õèìè÷åñêèå ñâîéñòâà, ôîðìà ìîëåêóë, ëîêàëèçàöèÿ è ïðîèñõîæäåíèå, àìèíîêèñëîòíûé ñîñòàâ), óæå íå îòâå÷àåò ïîëíîñòüþ âîçðîñøåìó óðîâíþ çíàíèé î èõ ñòðóêòóðå è ôóíêöèÿõ. Èç îãðîìíîãî êîëè÷åñòâà ïðèðîäíûõ áåëêîâ ñòðóêòóðà è ôóíêöèè ðàñøèôðîâàíû äëÿ îòíîñèòåëüíî íåáîëüøîãî ÷èñëà (íå áîëåå íåñêîëüêèõ ñîòåí), è ïîýòîìó ñòðóêòóðà è ôóíêöèè áåëêîâ ïîêà íå ìîãóò ñëóæèòü îñíîâîé äëÿ èõ ðàöèîíàëüíîé êëàññèôèêàöèè. Ïîæàëóé, òîëüêî äëÿ îäíîé ãðóïïû áåëêîâ, îáëàäàþùèõ ñïîñîáíîñòüþ êàòàëèçèðîâàòü õèìè÷åñêèå ðåàêöèè, ò.å. ôåðìåíòîâ, ðàçðàáîòàíà ñòðîéíàÿ ñèñòåìà íîìåíêëàòóðû è êëàññèôèêàöèè, â îñíîâó êîòîðîé ïîëîæåíû òèïû êàòàëè-çèðóåìûõ õèìè÷åñêèõ ðåàêöèé è õèìè÷åñêàÿ ïðèðîäà ðåàãèðóþùèõ âåùåñòâ. Îäíàêî ïîëíîñòüþ èäåíòèôèöèðîâàííûå äî ñèõ ïîð ôåðìåíòû òàêæå ñîñòàâëÿþò íåçíà÷èòåëüíóþ äîëþ áåëêîâ (ôåðìåíòîâ îïèñàíî áîëåå 3000). Òåì íå ìåíåå ôóíêöèîíàëüíûé ïðèíöèï, ðåêîìåíäóåìûé íåêîòîðûìè àâòîðàìè, õîòÿ è íå ìîæåò ñëóæèòü óíèâåðñàëüíîé îñíîâîé äëÿ êëàññèôèêàöèè âñåõ áåëêîâ, ïðåäñòàâëÿåò îïðåäåëåííûé èíòåðåñ. Â ñîîòâåòñòâèè ñ ôóíêöèîíàëüíûì ïðèíöèïîì ðàçëè÷àþò 12 ãëàâíûõ êëàññîâ áåëêîâ: 1) êàòàëèòè÷åñêè àêòèâíûå áåëêè (ôåðìåíòû); 2) áåëêè-ãîðìîíû (õîòÿ åñòü è ñòåðîèäíûå ãîðìîíû); 3) áåëêè-ðåãóëÿòîðû àêòèâíîñòè ãåíîìà; 4) çàùèòíûå áåëêè (àíòèòåëà, áåëêè ñâåðòûâàþùåé è àíòèñâåðòûâàþùåé ñèñòåì êðîâè); 5) òîêñè÷åñêèå áåëêè; 6) òðàíñïîðòíûå áåëêè; 7) ìåìáðàííûå áåëêè; 8) ñîêðàòèòåëüíûå áåëêè; 9) ðåöåïòîðíûå áåëêè; 10) áåëêè-èíãèáèòîðû ôåðìåíòîâ; 11) áåëêè âèðóñíîé îáîëî÷êè; 12) áåëêè ñ èíûìè ôóíêöèÿìè. Áûëè ïðåäïðèíÿòû òàêæå ïîïûòêè êëàññèôèöèðîâàòü áåëêè, èñõîäÿ èç îñîáåííîñòåé âòîðè÷íîé è òðåòè÷íîé ñòðóêòóðû. Â ñîîòâåòñòâèè ñ ýòèì ðàçëè÷àþò à-, â-, à+â- è à/â-áåëêè. à-Áåëêè ñîäåðæàò òîëüêî à-ñïèðàëè (íå ìåíåå 60%), â-áåëêè — òîëüêî â-ñòðóêòóðû (íå ìåíåå äâóõ àíòèïàðàëëåëüíûõ öåïåé), à+â-áåëêè — òå è äðóãèå ñòðóêòóðû â ïðåäåëàõ îäíîé ïîëèïåïòèäíîé öåïè (ïðèìåð—ìîëåêóëû ëèçîöèìà), à êëàññ à/â-áåëêîâ ñîäåðæèò ìíîæåñòâî à- è â-ñòðóêòóð, ÷åðåäóþùèõñÿ âäîëü ïîëèïåïòèäíîé öåïè èëè äîìåíà (ñì. ðèñ. 1.19). Äîìåíû ñîçäàþòñÿ îáúåäèíåíèåì è ÷åðåäîâàíèåì à-ñïèðàëåé è â-ñëîåâ, ìåæäó êîòîðûìè îòêðûâàþòñÿ áîëåå ðûõëûå ñòðóêòóðû. Ó÷èòûâàÿ íåîáõîäèìîñòü äëÿ ñòóäåíòà-ìåäèêà îñíîâàòåëüíîãî çíàêîìñòâà ñ îòäåëüíûìè ãðóïïàìè áåëêîâ, êîòîðûå ìîãóò ñëóæèòü ïðåäìåòîì èçó÷åíèÿ â åãî áóäóùåé ïðîôåññèîíàëüíîé äåÿòåëüíîñòè (íàïðèìåð, áåëêè êðîâè), ïðèâîäèì ñòàðóþ êëàññèôèêàöèþ áåëêîâ ñ êðàòêîé õàðàêòåðèñòèêîé íîâûõ äàííûõ î ñòðóêòóðå, ñîñòàâå è ñâîéñòâàõ îòäåëüíûõ ïðåäñòàâèòåëåé. Ñîãëàñíî ýòîé êëàññèôèêàöèè, îáøèðíûé êëàññ áåëêîâûõ âåùåñòâ â çàâèñèìîñòè îò õèìè÷åñêîãî ñîñòàâà äåëÿò íà ïðîñòûå è ñëîæíûå áåëêè . Ïðîñòûå áåëêè ïîñòðîåíû èç îñòàòêîâ àìèíîêèñëîò è ïðè ãèäðîëèçå ðàñïàäàþòñÿ ñîîòâåòñòâåííî òîëüêî íà ñâîáîäíûå àìèíîêèñëîòû. Ñëîæíûå áåëêè—ýòî äâóõêîìïîíåíòíûå áåëêè, êîòîðûå ñîñòîÿò èç êàêîãî-ëèáî ïðîñòîãî áåëêà è íåáåëêîâîãî êîìïîíåíòà, íàçûâàåìîãî ïðî- ñòåòè÷åñêîé ãðóïïîé. Ïðè ãèäðîëèçå ñëîæíûõ áåëêîâ, ïîìèìî ñâîáîäíûõ àìèíîêèñëîò, îñâîáîæäàåòñÿ íåáåëêîâàÿ ÷àñòü èëè ïðîäóêòû åå ðàñïàäà. Ïðîñòûå áåëêè â ñâîþ î÷åðåäü äåëÿòñÿ íà îñíîâàíèè íåêîòîðûõ óñëîâíî âûáðàííûõ êðèòåðèåâ íà ðÿä ïîäãðóïï: ïðîòàìèíû, ãèñòîíû, àëüáóìèíû, ãëîáóëèíû, ïðîëàìèíû, ãëþòåëèíû è äð. Êëàññèôèêàöèÿ ñëîæíûõ áåëêîâ (ñì. ãëàâó 2) îñíîâàíà íà õèìè÷åñêîé ïðèðîäå âõîäÿùåãî â èõ ñîñòàâ íåáåëêîâîãî êîìïîíåíòà. Â ñîîòâåòñòâèè ñ ýòèì ðàçëè÷àþò ôîñôîïðîòåèíû (ñîäåðæàò ôîñôîðíóþ êèñëîòó), õðîìîïðîòåèíû (â ñîñòàâ èõ âõîäÿò ïèãìåíòû), íóêëåîïðîòåèíû (ñîäåðæàò íóêëåèíîâûå êèñëîòû), ãëèêîïðîòåèíû (ñîäåðæàò óãëåâîäû), ëèïîïðîòåèíû (ñîäåðæàò ëèïèäû) è ìåòàëëîïðîòåèíû (ñîäåðæàò ìåòàëëû). ÕÈÌÈß ÏÐÎÑÒÛÕ ÁÅËÊÎÂ Ïðîòàìèíû è ãèñòîíû. Äàííàÿ ãðóïïà áåëêîâ îòëè÷àåòñÿ ðÿäîì õàðàêòåðíûõ ôèçèêî-õèìè÷åñêèõ ñâîéñòâ, ñâîåîáðàçèåì àìèíîêèñëîòíîãî ñîñòàâà è ïðåäñòàâëåíà â îñíîâíîì áåëêàìè ñ íåáîëüøîé ìîëåêóëÿðíîé ìàññîé. Ïðîòàìèíû îáëàäàþò âûðàæåííûìè îñíîâíûìè ñâîéñòâàìè, îáóñëîâëåííûìè íàëè÷èåì â èõ ñîñòàâå îò 60 äî 85% àðãèíèíà. Òàê, ñàëüìèí, âûäåëåííûé èç ìîëîê ñåìãè, ñîñòîèò íà 85% èç àðãèíèíà. Âûñîêèì ñîäåðæàíèåì àðãèíèíà îòëè÷àåòñÿ äðóãîé õîðîøî èçó÷åííûé áåëîê—êëó- ïåèí, âûäåëåííûé èç ìîëîê ñåëüäè: èç 30 àìèíîêèñëîò â íåì íà äîëþ àðãèíèíà ïðèõîäèòñÿ 21 îñòàòîê. Ðàñøèôðîâàíà ïåðâè÷íàÿ ñòðóêòóðà êëó- ïåèíà. Ïðîòàìèíû õîðîøî ðàñòâîðèìû â âîäå, èçîýëåêòðè÷åñêàÿ òî÷êà èõ âîäíûõ ðàñòâîðîâ íàõîäèòñÿ â ùåëî÷íîé ñðåäå. Ïî ñîâðåìåííûì ïðåäñòàâëåíèÿì, ïðîòàìèíû ñêîðåå âñåãî ÿâëÿþòñÿ ïåïòèäàìè, à íå áåëêàìè, ïîñêîëüêó èõ ìîëåêóëÿðíàÿ ìàññà íå ïðåâûøàåò 5000. Îíè ñîñòàâëÿþò áåëêîâûé êîìïîíåíò â ñòðóêòóðå ðÿäà ñëîæíûõ áåëêîâ. Ãèñòîíû òàêæå ÿâëÿþòñÿ áåëêàìè îñíîâíîãî õàðàêòåðà. Â èõ ñîñòàâ âõîäÿò ëèçèí è àðãèíèí, ñîäåðæàíèå êîòîðûõ, îäíàêî, íå ïðåâûøàåò 20—30%. Ìîëåêóëÿðíàÿ ìàññà ãèñòîíîâ íàìíîãî áîëüøå íèæíåãî ïðåäåëà ìîëåêóëÿðíîé ìàññû áåëêîâ. Ýòè áåëêè ñîñðåäîòî÷åíû â îñíîâíîì â ÿäðàõ êëåòîê â ñîñòàâå äåçîêñèðèáîíóêëåîïðîòåèíîâ è èãðàþò âàæíóþ ðîëü â ðåãóëÿöèè ýêñïðåññèè ãåíîâ (ñì. ãëàâû 2 è 3). Ïðîëàìèíû è ãëþòåëèíû. Ýòî áåëêè ðàñòèòåëüíîãî ïðîèñõîæäåíèÿ, îòëè÷àþòñÿ ñâîåîáðàçèåì àìèíîêèñëîòíîãî ñîñòàâà è ôèçèêî-õèìè÷åñêèõ ñâîéñòâ. Îíè ñîäåðæàòñÿ â îñíîâíîì â ñåìåíàõ çëàêîâ (ïøåíèöà, ðîæü, ÿ÷ìåíü è äð.), ñîñòàâëÿÿ îñíîâíóþ ìàññó êëåéêîâèíû. Õàðàêòåðíîé îñîáåí-íîñòüþ ïðîëàìèíîâ ÿâëÿåòñÿ ðàñòâîðèìîñòü â 60—80% âîäíîì ðàñòâîðå ýòàíîëà, â òî âðåìÿ êàê âñå îñòàëüíûå ïðîñòûå áåëêè â ýòèõ óñëîâèÿõ îáû÷íî âûïàäàþò â îñàäîê. Íàèáîëåå èçó÷åíû îðèçåíèí (èç ðèñà), ãëþòå- íèí è ãëèàäèí (èç ïøåíèöû), çåèí (èç êóêóðóçû), ãîðäåèí (èç ÿ÷ìåíÿ) è äð. Óñòàíîâëåíî, ÷òî ïðîëàìèíû ñîäåðæàò 20—25% ãëóòàìèíîâîé êèñëîòû è 10—15% ïðîëèíà. Àëüáóìèíû è ãëîáóëèíû. Ýòè áåëêè îòíîñÿòñÿ ê áåëêàì, øèðîêî ðàñïðîñòðàíåííûì â îðãàíàõ è òêàíÿõ æèâîòíûõ. Íàèáîëåå áîãàòû èìè áåëêè ñûâîðîòêè êðîâè, ìîëîêà, ÿè÷íûé áåëîê, ìûøöû è äð. Â ïëàçìå êðîâè ÷åëîâåêà â íîðìå ñîäåðæèòñÿ îêîëî 7% áåëêîâ, ïðåäñòàâëåííûõ ïðåèìóùåñòâåííî àëüáóìèíàìè è ãëîáóëèíàìè. Àëüáóìèíû è ãëîáóëèíû—ýòî ãëîáóëÿðíûå áåëêè, ðàçëè÷àþùèåñÿ ïî ðàñòâîðèìîñòè (òàáë. 1.6). Íåîáõîäèìî îòìåòèòü, ÷òî ñàìî îïðåäåëåíèå «àëüáóìèíû» è «ãëîáóëèíû» îñíîâàíî íà èõ ðàñòâîðèìîñòè â äèñòèëëèðîâàííîé âîäå è ïîëóíàñû- Òàáëèöà 1.6. Ðàñòâîðèìîñòü àëüáóìèíîâ è ãëîáóëèíîâ Ðàñòâîðèòåëü Àëüáóìèíû Ãëîáóëèíû Äèñòèëëèðîâàííàÿ âîäà Ñëàáûå ñîëåâûå ðàñòâîðû ¹Ñ1 Íàñûùåííûé ðàñòâîð Êà,804 Íàñûùåííûé ðàñòâîð ÊàÑ1 Ïîëóíàñûùåííûé ðàñòâîð (ÊÍ4)28 04 Íàñûùåííûé ðàñòâîð (ÊÍ4)2804 Ðàñòâîðèìû » » » » Íåðàñòâîðèìû Íåðàñòâîðèìû Ðàñòâîðèìû Íåðàñòâîðèìû » » » ùåííîì ðàñòâîðå (ÍÈ4)2804. Îäíàêî, êàê ïîêàçûâàþò äàííûå òàáë. 1.6, ãëîáóëèíû ðàñòâîðèìû òîëüêî â ðàçáàâëåííûõ ñîëåâûõ ðàñòâîðàõ. Ðàçëè÷íóþ ðàñòâîðèìîñòü àëüáóìèíîâ è ãëîáóëèíîâ ñûâîðîòêè êðîâè ðàíüøå øèðîêî èñïîëüçîâàëè â êëèíè÷åñêîé ïðàêòèêå äëÿ èõ ôðàêöèîíèðîâàíèÿ è êîëè÷åñòâåííîãî îïðåäåëåíèÿ (ñì. ãëàâó 17). Â íàñòîÿùåå âðåìÿ êà÷åñòâåííûé ñîñòàâ è ñîäåðæàíèå ñûâîðîòî÷íûõ áåëêîâ îïðåäåëÿþò ñ ïîìîùüþ ýëåêòðîôîðåçà íà áóìàãå è â ïîëèàêðèëàìèäíîì ãåëå â íåáîëüøîì êîëè÷åñòâå ñûâîðîòêè êðîâè. Òèïè÷íàÿ ýëåêòðî- ôîðåãðàììà áåëêîâ ñûâîðîòêè êðîâè, à òàêæå ñîîòíîøåíèå îòäåëüíûõ ôðàêöèé ïðåäñòàâëåíû â ãëàâå 17. Àëüáóìèíû è ãëîáóëèíû îòëè÷àþòñÿ äðóã îò äðóãà òàêæå ïî ìîëåêóëÿðíîé ìàññå — ñîîòâåòñòâåííî 40000—70000 è 150000 è áîëåå. Èç ñûâîðîòêè êðîâè íå òîëüêî âûäåëåí àëüáóìèí â ÷èñòîì âèäå, íî è îïðåäåëåíà ïåðâè÷íàÿ ñòðóêòóðà åãî åäèíñòâåííîé ïîëèïåïòèäíîé öåïè (575 àìèíîêèñëîòíûõ îñòàòêîâ). Àëüáóìèí èìååò îòíîñèòåëüíî íèçêóþ èçîýëåêòðè÷åñêóþ òî÷êó (4,7) è âûñîêèé îòðèöàòåëüíûé çàðÿä ïðè ðÍ 8,6, áëàãîäàðÿ ÷åìó îí ìèãðèðóåò ñ áîëüøîé ñêîðîñòüþ â ýëåêòðè÷åñêîì ïîëå ê àíîäó. Ïðèíÿòî ñ÷èòàòü, ÷òî ïðèìåðíî 75—80% îñìîòè÷åñêîãî äàâëåíèÿ áåëêîâ ñûâîðîòêè êðîâè ïðèõîäèòñÿ íà àëüáóìèíû; êðîìå òîãî, îñíîâíîé ôóíêöèåé èõ ñ÷èòàþò òðàíñïîðò æèðíûõ êèñëîò. Îäíàêî òî÷íàÿ ôóíêöèÿ àëüáóìèíîâ íå ñîâñåì ÿñíà. Èçâåñòíû ñëó÷àè, êîãäà ó íåêîòîðûõ ëþäåé â êðîâè ôàêòè÷åñêè îòñóòñòâóþò àëüáóìèíû (âðîæäåííàÿ àíîìàëèÿ), íî îíè ïðàêòè÷åñêè çäîðîâû. Ãëîáóëèíû, ïðåäñòàâëåííûå à1-ôðàêöèåé, ñîäåðæàòñÿ â êðîâè â êîìïëåêñå ñ áèëèðóáèíîì è ñ ëèïîïðîòåèíàìè âûñîêîé ïëîòíîñòè. Ãëîáóëèíû, ìèãðèðóþùèå ïðè ýëåêòðîôîðåçå â âèäå à2-ôðàêöèè, ñîäåðæàò ãëîáóëèí è íåèçâåñòíûé ãëèêîïðîòåèí. â-Ãëîáóëèíû âêëþ÷àþò ðÿä âàæíûõ â ôóíêöèîíàëüíîì îòíîøåíèè áåëêîâ, â ÷àñòíîñòè òðàíñôåððèí—áåëîê, îòâåòñòâåííûé çà òðàíñïîðò æåëåçà. Ñ ýòîé æå ôðàêöèåé ñâÿçàí öåðóëîïëàçìèí — áåëîê, òðàíñïîðòèðóþùèé èîíû ìåäè. Îòñóòñòâèå ýòîãî áåëêà ïðèâîäèò ê ðàçâèòèþ ãåïàòîöåðåáðàëüíîé äèñòðîôèè, ïðè êîòîðîé íàáëþäàåòñÿ îòðàâëåíèå îðãàíèçìà èîíàìè ñâîáîäíîé ìåäè. Â îñíîâå áîëåçíè ëåæèò âðîæäåííûé äåôèöèò ñèíòåçà öåðóëîïëàçìèíà. Íàêîíåö, âî ôðàêöèè â- ãëîáóëèíîâ ñîäåðæèòñÿ ïðîòðîìáèí, ÿâëÿþùèéñÿ ïðåäøåñòâåííèêîì òðîìáèíà —áåëêà, îòâåòñòâåííîãî çà ïðåâðàùåíèå ôèáðèíîãåíà êðîâè â ôèáðèí ïðè ñâåðòûâàíèè êðîâè. Ôðàêöèÿ ó-ãëîáóëèíîâ ÿâëÿåòñÿ íàèáîëåå ãåòåðîãåííîé. Èçâåñòíî ìíîæåñòâî àíòèòåë, ðàçëè÷àþùèõñÿ ïåðâè÷íîé ñòðóêòóðîé. Ýëåêòðîôîðåòè÷å- ñêè îíè îòêðûâàþòñÿ ãëàâíûì îáðàçîì â ó-ãëîáóëèíîâîé è ÷àñòè÷íî â â2-ãëîáóëèíîâîé ôðàêöèÿõ. Ñòðóêòóðà è ôóíêöèÿ ó-ãëîáóëèíîâ áîëåå ïîäðîáíî ðàññìîòðåíû äàëåå (ñì. ãëàâó 2, «Ãëèêîïðîòåèíû»). ÏÐÈÐÎÄÍÛÅ ÏÅÏÒÈÄÛ Â ïîñëåäíèå ãîäû çíà÷èòåëüíî ïîâûñèëñÿ èíòåðåñ ê ñòðóêòóðå è ôóíêöèÿì âñòðå÷àþùèõñÿ â ñâîáîäíîì ñîñòîÿíèè â îðãàíèçìå íèçêîìîëåêóëÿðíûõ ïåïòèäîâ, âûïîëíÿþùèõ ðÿä ñïåöèôè÷åñêèõ áèîëîãè÷åñêèõ ôóíêöèé. Êîðîòêèå ïåïòèäû, ñîäåðæàùèå äî 10 àìèíîêèñëîò, ïðèíÿòî íàçûâàòü îëèãîïåïòèäàìè; â òî æå âðåìÿ ïîëèïåïòèäû è áåëêè ñ÷èòàþòñÿ âçàèìîçàìåíÿåìûìè, õîòÿ òåðìèíîì «ïîëèïåïòèäû» ÷àùå îáîçíà÷àþò ïðîäóêòû ñ ìîë. ì. ìåíåå 10000. Â íåêîòîðûõ áèîàêòèâíûõ ïåïòèäàõ èìåþòñÿ íåîáû÷íûå àìèíîêèñëîòû, íå âñòðå÷àþùèåñÿ â ïðèðîäíûõ áåëêàõ, èëè ïðîèçâîäíûå îáû÷íûõ àìèíîêèñëîò (ãîðìîíû, àíòèáèîòèêè). Ìíåíèå î òîì, ÷òî ïåïòèäû ìîãóò èãðàòü ðîëü ïðîìåæóòî÷íûõ ïðîäóêòîâ íà ïóòè ñèíòåçà áåëêà, íå ïîäòâåðäèëîñü, ïîñêîëüêó, êàê ïîêàçàíî â ãëàâå 14, ýòîò ïðîöåññ âî âñåõ êëåòêàõ ó âñåõ æèâûõ îðãàíèçìîâ îñóùåñòâëÿåòñÿ ¸å ïîóî ìàòðè÷íûì ïóòåì. Ïðèðîäíûå ïåïòèäû, íàäåëåííûå áèîëîãè÷åñêîé àêòèâíîñòüþ, â çàâèñèìîñòè îò õàðàêòåðà äåéñòâèÿ è ïðîèñõîæäåíèÿ ïðèíÿòî äåëèòü íà 4 ãðóïïû: ïåïòèäû, îáëàäàþùèå ãîðìîíàëüíîé àêòèâíîñòüþ (âàçîïðåññèí, îêñè- òîöèí, êîðòèêîòðîïèí, ãëþêàãîí, êàëüöèòîíèí, ìåëàíîöèòñòèìóëèðóþùèé ãîðìîí, ðèëèçèíã-ôàêòîðû ãèïîòàëàìóñà è äð.; ñì. ãëàâó 8); 2) ïåïòèäû, ïðèíèìàþùèå ó÷àñòèå â ïðîöåññå ïèùåâàðåíèÿ (â ÷àñòíîñòè, ãàñòðèí è ñåêðåòèí; ñì. ãëàâó 12); 3) ïåïòèäû, èñòî÷íèê êîòîðûõ—à2-ãëîáóëèíîâàÿ ôðàêöèÿ ñûâîðîòêè êðîâè (òàêèå, êàê àíãèîòåíçèí, áðàäèêèíèí è êàëëèäèí); 4) íåéðîïåïòèäû. Â ïîñëåäíåå âðåìÿ âûÿñíåíû íåêîòîðûå çàêîíîìåðíîñòè ñèíòåçà ôèçèîëîãè÷åñêè àêòèâíûõ ïåïòèäîâ èç áèîëîãè÷åñêè èíåðòíûõ ïðåäøåñòâåííèêîâ — áåëêîâ â ðåçóëüòàòå ïðîöåññà, íàçûâàåìîãî ïîñòòðàíñëÿöèîííîé ìîäèôèêàöèåé (ïîñòñèíòåòè÷åñêèå ïðåâðàùåíèÿ áåëêîâîé ìîëåêóëû). Èçâåñòíî, íàïðèìåð, ÷òî àíãèîòåíçèíû (ïðåäñòàâëåííûå îêòàïåïòèäàìè), îêàçûâàþùèå âûðàæåííîå ñîñóäîñóæèâàþùåå äåéñòâèå, îáðàçóþòñÿ èç ïðèñóòñòâóþùåãî â ñûâîðîòêå êðîâè íåàêòèâíîãî áåëêà àíãèîòåíçèíîãåíà â ðåçóëüòàòå ïîñëåäîâàòåëüíîãî äåéñòâèÿ ðÿäà ïðîòåîëèòè÷åñêèõ ôåðìåíòîâ (ðåíèíà è îñîáîãî ôåðìåíòà, ó÷àñòâóþùåãî â ïðåâðàùåíèè íåàêòèâíîãî àíãèîòåíçèíà I â àêòèâíûé àíãèîòåíçèí II). ÀÍÃÈÎÒÅÍÇÈÍÎÃÅÍ (íåàêòèâíûé) Àñï-Àðã-Âàë—Òèð-Èëå-Ã èñ—Ïðî-Ôåí-Ãèñ-Ëåé-Âàë—Òèð-Ñåð-ïîëèïåïòèä I Ðåíèí 1 ÀÍÃÈÎÒÅÍÇÈÍ I (íåàêòèâíûé) Àñï—Àðã—Âàë—Òèð—Èëå—Ã èñ—Ïðî— Ôåí—Ã èñ—Ëåé I Êàðáîêñè- ^ X êàòåïñèí ÀÍÃÈÎÒÅÍÇÈÍ II (àêòèâíûé) Àñï-Àðã-Âàë-Ò èð-Èëå-Ã èñ-Ïðî-Ôåí Ê ãðóïïå âàçîàêòèâíûõ (îêàçûâàþùèõ âëèÿíèå íà òîíóñ ñîñóäîâ) ïåïòèäîâ îòíîñÿòñÿ, êðîìå òîãî, øèðîêî ïðèìåíÿåìûå â ìåäèöèíñêîé ïðàêòèêå áðàäèêèíèí è êàëëèäèí. Áðàäèêèíèí ïðåäñòàâëÿåò ñîáîé íîíàïåïòèä: Í-Àðã-Ïðî-Ïðî-Ãëè-Ôåí-Ñåð-Ïðî-Ôåí-Àðã-ÎÍ. Êàëëèäèí ïðåäñòàâëåí äåêàïåïòèäîì, îáðàçóþùèìñÿ èç íåàêòèâíîãî ïëàçìåííîãî áåëêà êèíèíîãåíà, è îòëè÷àåòñÿ îò áðàäèêèíèíà ïðèñóòñòâèåì íà Îêîíöå åùå îäíîãî àìèíîêèñëîòíîãî îñòàòêà (Ëèç): Í-Ëèç-Àðã-Ïðî-Ïðî-Ãëè-Ôåí-Ñåð-Ïðî-Ôåí-Àðã-ÎÍ. Ñîâñåì íåäàâíî èç ýêñòðàêòîâ òêàíè ïðåäñåðäèÿ (íî íå èç æåëóäî÷êîâ ñåðäöà) ÷åëîâåêà è æèâîòíûõ áûëè âûäåëåíû áèîëîãè÷åñêè àêòèâíûå ïåïòèäû, ðåãóëèðóþùèå òîíóñ ñîñóäèñòîé ñèñòåìû è ýëåêòðîëèòíûé îáìåí. Ôèçèîëîãè÷åñêèé ýôôåêò èõ îêàçàëñÿ ïðîòèâîïîëîæíûì âëèÿíèþ ñèñòåìû ðåíèí—àíãèîòåíçèí—àëüäîñòåðîí. Îí âûðàæàåòñÿ â ñîñóäîðàñøèðÿþùåì äåéñòâèè, óñèëåíèè êëóáî÷êîâîé ôèëüòðàöèè è ñòèìóëÿöèè âûâåäåíèÿ íàòðèÿ è õëîðèäîâ çà ñ÷åò óãíåòåíèÿ èõ ðåàáñîðáöèè â êàíàëüöàõ. Ýòè ïåïòèäû ïîëó÷èëè íàçâàíèå àòðèîïåïòèäîâ (îò ëàò. à1ïî — ïðåäñåðäèå). Îíè ïîñòðîåíû èç ðàçíîãî ÷èñëà àìèíîêèñëîò (îò 23 äî 100), íî îáÿçàòåëüíûì óñëîâèåì äëÿ ïðîÿâëåíèÿ áèîëîãè÷åñêîãî ýôôåêòà ÿâëÿåòñÿ íàëè÷èå â ìîëåêóëå 17-÷ëåííîé êîëüöåâîé ñòðóêòóðû, îáðàçóþùåéñÿ çà ñ÷åò äèñóëü- ôèäíîé ñâÿçè ìåæäó îñòàòêàìè öèñòåèíà. Âíóòðèêëåòî÷íûì ïîñðåäíèêîì äåéñòâèÿ àòðèîïåïòèäîâ îêàçàëñÿ öèêëè÷åñêèé ãóàíîçèíìîíîôîñôàò (öÃÌÔ), ñèíòåç êîòîðîãî îñóùåñòâëÿåòñÿ â ðåçóëüòàòå àêòèâèðîâàíèÿ ìåìáðàííîãî ôåðìåíòà ãóàíèëàòöèêëàçû; äåéñòâèå àäåíèëàòöèêëàçû, íàïðîòèâ, òîðìîçèòñÿ ïîä âëèÿíèåì àòðèîïåï- òèäîâ. Âî âñåõ æèâîòíûõ òêàíÿõ è â íåêîòîðûõ ðàñòåíèÿõ øèðîêî ðàñïðîñòðàíåí íèçêîìîëåêóëÿðíûé òðèïåïòèä ãëóòàòèîí, ôóíêöèè êîòîðîãî ïîêà íå âûÿñíåíû äîñòàòî÷íî ïîëíî, õîòÿ îí îòêðûò ñðàâíèòåëüíî äàâíî. Ãëóòà- òèîí ïðåäñòàâëÿåò ñîáîé àòèïè÷íûé òðèïåïòèä (â êîòîðîì â îáðàçîâàíèè îäíîé èç ïåïòèäíûõ ñâÿçåé ó÷àñòâóåò íå à-êàðáîêñèëüíàÿ, à ó-êàðáîê- ñèëüíàÿ ãðóïïà ãëóòàìàòà) ñëåäóþùåãî ñòðîåíèÿ: ó-ãëóòàìèë-öèñòåèíèë- ãëèöèí: ÍÎÎÑ-ÑÍ-ÑÍ2-ÑÍ2-ÑÎ-ÌÍ-ÑÍ-ÑÍ2 _8Í Ãëóòàòèîí (âîññòàíîâëåííûé) Öèñòåèí ÿâëÿåòñÿ ñîñòàâíîé ÷àñòüþ ãëóòàòèîíà, ïîýòîìó ïîñëåäíèé ìîæåò íàõîäèòüñÿ â âîññòàíîâëåííîé (8Í) è â îêèñëåííîé (8-8) ôîðìàõ (ñîêðàùåííî îáîçíà÷àþòñÿ Ã-8Í è Ã-8-8-Ã), ÷òî, ïî-âèäèìîìó, èìååò îòíîøåíèå ê áèîëîãè÷åñêîé ðîëè ãëóòàòèîíà â îðãàíèçìå. Èíòåðåñ ê ïðèðîäíûì ïåïòèäàì â çíà÷èòåëüíîé ñòåïåíè îáóñëîâëåí íåîáû÷íî âûñîêîé èõ áèîëîãè÷åñêîé àêòèâíîñòüþ. Îíè îêàçûâàþò ìîùíîå ôàðìàêîëîãè÷åñêîå äåéñòâèå íà ìíîæåñòâî ôèçèîëîãè÷åñêèõ ôóíêöèé îðãàíèçìà. Â òî æå âðåìÿ áûëè çàìå÷åíû íèçêàÿ ñòàáèëüíîñòü è áûñòðûé ðàñïàä èõ â îðãàíèçìå ïðè ôèçèîëîãè÷åñêèõ çíà÷åíèÿõ ðÍ ñðåäû. Âñå ýòî ñïîñîáñòâîâàëî ðàçâèòèþ èññëåäîâàíèé êàê â îáëàñòè ïðåïàðàòèâíîãî âûäåëåíèÿ ïðèðîäíûõ ïåïòèäîâ èç îðãàíîâ è òêàíåé (âêëþ÷àÿ ïîëó÷åíèå áèîëîãè÷åñêè àêòèâíûõ ïåïòèäîâ èç ïðåäøåñòâåííèêîâ ìåòîäàìè îãðàíè÷åííîãî ïðîòåîëèçà ðÿäà õîðîøî èçâåñòíûõ ãîðìîíîâ), òàê è â îáëàñòè õèìè÷åñêîãî ñèíòåçà. Ïîëó÷åíèå ðÿäà áèîëîãè÷åñêè àêòèâíûõ íåéðîïåïòèäîâ èç ãîðìîíîâ ãèïîôèçà, â ÷àñòíîñòè ýíäîðôèíîâ è ýíêåôàëèíîâ, íàäåëåííûõ ìîùíûì îáåçáîëèâàþùèì äåéñòâèåì (ïóòåì ñâÿçûâàíèÿ ðåöåïòîðîâ îïðåäåëåííûõ êëåòîê ìîçãà), â ñîòíè è òûñÿ÷è ðàç ïðåâîñõîäÿùèì àíàëãåçèðóþùèé ýôôåêò ìîðôèíà, îïèñàíî â ãëàâå 8. Èç òêàíè ìîçãà âûäåëåí òàêæå 6-ïåïòèä ñíà; ðÿä äðóãèõ íåéðîïåïòèäîâ ïðèíèìàåò ó÷àñòèå â áèîõèìè÷åñêèõ ìåõàíèçìàõ ïàìÿòè, ñòðàõà, îáó÷åíèÿ è ò.ä. Äëÿ ïîâûøåíèÿ ñòàáèëüíîñòè ïåïòèäîâ ïðè ââåäåíèè â îðãàíèçì ïðåäïðèíÿòû ïîïûòêè õèìè÷åñêîãî ñèíòåçà ïåïòèäîâ, â êîòîðûõ îäèí èëè íåñêîëüêî àìèíîêèñëîòíûõ îñòàòêîâ Ü-ðÿäà çàìåùàþò îñòàòêàìè Á- àìèíîêèñëîò. Ïîäîáíàÿ çàìåíà, íå âûçûâàÿ ñíèæåíèÿ áèîàêòèâíîñòè, çàùèùàåò ïåïòèä îò âîçäåéñòâèÿ ïðîòåèíàç òêàíåé, ñïîñîáñòâóÿ ïðîëîíãèðîâàíèþ ýôôåêòà ïðåïàðàòà. Ñðåäè åñòåñòâåííî âñòðå÷àþùèõñÿ íåáîëüøèõ ïåïòèäîâ ñëåäóåò óêàçàòü íà àíòèáèîòèê ãðàìèöèäèí 8, âûäåëåííûé èç ÁàåØèç ÜãåÓ18 è ïðåäñòàâëÿþùèé ñîáîé öèêëè÷åñêèé äåêàïåïòèä: ôåí —¦ I Ïðî <— 1--Ëåé —¦ 1,-Îðí —> è-Âàë —> Ü-Ïðî Âàë <— 1_-Îðí <— 1_-Ëåé *— Ð-Ôåí Ã ðàìèöèäèí 5 Êàê âèäíî, â ñòðóêòóðå ãðàìèöèäèíà 8 èìåþòñÿ 2 îñòàòêà îðíèòèíà (Îðí), ïðîèçâîäíûå àìèíîêèñëîòû àðãèíèíà è 2 îñòàòêà íåïðèðîäíûõ Á-èçîìåðîâ ôåíèëàëàíèíà. Ñòðåëêè óêàçûâàþò íàïðàâëåíèå ñèíòåçà îò ÍÍ2-ãðóïï ê ÑÎÎÍ-ãðóïïàì êàæäîãî îñòàòêà, è âñëåäñòâèå öèêëè÷íîñòè ãðàìèöèäèí 8 íå èìååò êîíöà. Øèðîêîå ïðèìåíåíèå, îñîáåííî â ïèùåâîé ïðîìûøëåííîñòè, â êà÷åñòâå çàìåíèòåëÿ ñàõàðà ïîëó÷èë èñêóññòâåííûé (ãåííîèíæåíåðíûé ñèíòåç) äèïåïòèä, ñîñòîÿùèé èç Ü-èçîìåðîâ àñïàðàãèíîâîé êèñëîòû è ìåòèëîâîãî ýôèðà ôåíèëàëàíèíà, íàçâàííûé àñïàðòàìîì: íîîñ-ñí2-ñí\|ñî-ìí òÑÍ-ÑÍã Ëíã' ÑÎ-ÎÑÍç Àñïàðòàì Àñïàðòàì â ñîòíè ðàç ñëàùå ñàõàðà è ëåãêî ðàñïàäàåòñÿ â îðãàíèçìå íà äâå ñâîáîäíûå àìèíîêèñëîòû, àáñîëþòíî áåçâðåäíûå äëÿ îðãàíèçìà; ïîýòîìó îí ðåêîìåíäîâàí â êà÷åñòâå çàìåíèòåëÿ ñàõàðà áîëüíûì äèàáåòîì. Ýòî ïðèìåð ïåïòèäà, íàäåëåííîãî îãðîìíûì áèîëîãè÷åñêèì ýôôåêòîì. << \| >> Èñòî÷íèê: Áåðåçîâ Ò. Ò., Êîðîâêèí Á. Ô.. Áèîëîãè÷åñêàÿ õèìèÿ: Ó÷åáíèê.— 3-å èçä., ïåðåðàá. è äîï.— Ì.: Ìåäèöèíà, 1998.— 704 ñ.. 1998 Åùå ïî òåìå Ãëàâà 1 ÕÈÌÈß ÁÅËÊÎÂ: 1. ãëàâàðåöåïöèÿ è ðåöåïòîðû ãîðìîíîâ 2. Ãëàâà 39 Ïàðåíòåðàëüíîå ïèòàíèå 3. ÃËÀÂÀ 2. ÎÏÈÑÀÍÈß ËÅÊÀÐÑÒÂÅÍÍÌÕ ÑÐÅÄÑÒÂ \|È ÄÅÈÑÒÂÓÞÙÈÕ ÂÅÙÅÑÒ 4. Ãëàâà 5 Ïîòðåáíîñòè îðãàíèçìà â áåëêå è ýíåðãèèÏîòðåáíîñòü â áåëêå 5. Ãëàâà 14 Ýíòåðàëüíîå è ïàðåíòåðàëüíîå ïèòàíèå 6. Ãëàâà 37 Ïèòàíèå â ïðîôèëàêòèêå è ëå÷åíèè îíêîëîãè÷åñêèõ è ãåìàòîëîãè÷åñêèõ áîëåçíåé 7. Ãëàâà 18Çàáîëåâàíèÿ ïîäæåëóäî÷íîé æåëåçû 8. ÃËÀÂÀ 2. ÎÏÈÑÀÍÈß ËÅÊÀÐÑÒÂÅÍÍÛÕ ÑÐÅÄÑÒ 9. Ãëàâà 11ÎÑÎÁÅÍÍÎÑÒÈ ÎÁÙÅÉ ÀÍÅÑÒÅÇÈÈ Â ÑÏÅÖÈÀËÜÍÛÕ ÎÁËÀÑÒßÕ ÕÈÐÓÐÃÈÈ 10. Ãëàâà 2. Ðàñïðåäåëåíèå, íàêîïëåíèå è ýëèìèíàöèÿ òîêñèíîâ 11. Ãëàâà 1 ÕÈÌÈß ÁÅËÊÎÂ 12. Ãëàâà 2ÕÈÌÈß ÑËÎÆÍÛÕ ÁÅËÊÎÂ 13. Ãëàâà 3ÕÈÌÈß ÍÓÊËÅÈÍÎÂÛÕ ÊÈÑËÎÒ 14. Ãëàâà 4 ÔÅÐÌÅÍÒÛ 15. Ãëàâà 8 ÃÎÐÌÎÍÛ 16. Ãëàâà 12 ÎÁÌÅÍ ÏÐÎÑÒÛÕ ÁÅËÊÎÂ
YZXStudio Load ZL1000 DSC_3640 Official specifications: I got it from ebay dealer f-t-2000, he is also known as Franky or 99centhobbies There exist a couple of different loads for usage with usb devices, this one is adjustable, can handle a lot of power and can control different fast charge schemes. DSC_3635 On the topside is a nearly noiseless fan and a multiturn trimpot to adjust the current with. DSC_3636 This side has a lot of electronic on it. The load is a IRF3710 mosfet transistor, controlled from a MCP6002 dual OpAmp. The trimpot has a TL431 reference across it. There is also two voltage regulators, one for the fan and one for the electronic. The "big" chip is a ATtiny24A microprocessor that controls the fast charge modes and keep an eye on maximum power level. led If you look closely on the board above, you will also find a small multicolored led, it has 3 leds inside. This led is used to signal different modes. DSC_3637 DSC_3638 DSC_3639 DSC_3641 Here it is together with a usb monitor that can show voltage, current, capacity and also usb coding. Load testing yzxStudio%20Load%20ZL1000%20voltage%20sweep I used the limit from the circuit board (i.e. 14 volt). At the currents the load is stable with any voltage, when the current increases the power is limited and there is some variation in the current. yzxStudio%20Load%20ZL1000%20voltage%20sweep%20min Here is a close look at the minimum current, it depend on the voltage (May be due to the fan) and it is not completely stable. yzxStudio%20Load%20ZL1000%20Load%20Test Can the load handle full power for some time? I decided to test for 30 minutes. The current looks fairly stable, but why do the power drops at the high power level? A look in the log file showed the voltage had dropped a bit, but that is not possible, except if my equipment is overloaded (Current limit was at 3A and I was only drawing 2.5A) Osc2 I put a current clamp around the wires and connected the scope. The current was above 3A sometimes. The circuit is oscillating. Osc4 Testing a bit more at different current and voltage levels shows that the circuit is very sensitive, sometimes it will oscillate, sometimes not. The difference can be a hand around the wires. I did try this with 3 different power supplies, with all of them there were oscillations. Temp2960 M1: 40,6°C, M2: 45,3°C, HS1: 54,1°C This IR photo is after 25 minutes with 12.5 watt load. Temp2961 M1: 39,6°C, HS1: 85,4°C Temp2962 M1: 61,7°C, M2: 51,3°C, HS1: 75,4°C This IR photo is after 25 minutes with 22 watt load. Temp2963 M1: 49,8°C, HS1: 111,8°C Everything is getting hot, this is the reason for the current change. yzxStudio%20Load%20ZL1000%20Load%20Test%20zoom Let’s take a look at how stable the load is, for this I have zoomed the current and power scales. I have about 0.1A drop in current, this is about 4%. The two spikes on the curves are because I touched the wires and affected the oscillations. Fast charge functions Using the two buttons on the circuit board it is possible to activate smart charge modes. QCtest It can select a specific QuickCharge voltage or cycle between them either fast or slow. Above I have selected a fast cycle. When the load is put into a QuickCharge charger the led will turn on and if one of the QC modes has been selected on the load it will switch the charger to that mode. It do not support the 20V QC mode. QC3test Selecting QuickCharge 3 mode it is possible to step the voltage up/down from a QC3 power supply. I pressed the +/- buttons a lot for this chart. PEup In MTKPE mode it can send up/down requests to the charger. Conclusion For testing usb chargers and power banks this load is very interesting, it can handle the power and it can select fast charge modes. The instability in the circuit can in some cases give misleading results. I am not very happy with the trimpot or the small buttons on the back. I would have like the buttons at the edge of the circuit board, one on each side, this would have been much easier to use without looking at them (Display is on the other side when connected to a monitor). The trimpot requires a tool to adjust. It is also a good idea to do all adjustments before starting a test, it gets rather hot! Even with the above issues I will call it a fairly good load and with way more functions than any other load on the market.
HomeLiving HealthyGeneral HealthVarious Factors Resulting In Repetitive Hiccups Various Factors Resulting In Repetitive Hiccups Introduction Hiccups are involuntary contractions of the diaphragm. Diaphragm is the muscle that separates the chest from your abdomen and plays a significant role in breathing. Each of this contraction is followed by sudden closure of the vocal cords that produces the characteristic “hic” sound. Your diaphragm is the muscle that lies just underneath your lungs. It denotes a boundary between your chest and abdomen . The role of the diaphragm is to regulate breathing . At the point when it relaxes, your lungs take in oxygen. At the point when it contracts , your lungs give out carbon dioxide. Some hiccups may last for a few minutes , though they can go on for even over 48 hours to a few days ion some cases. . Most instances of hiccups start and end suddenly. Symptoms of Hiccups Hiccupping is a symptom in itself. It might at times be joined by a slight tightening sensation in your chest, mid-region or throat. Hiccups are medically known as coordinated diaphragmatic shudder or singultus. They can happen independently or in sessions. When to See a Doctor Since hiccups are seldom a health-related crisis, you can wait and see if it goes away on its own . Different experts who might be associated with treating hiccups include an otolaryngologist, a gastroenterologist, a nervous system specialist, a pulmonologist, or a general medicine physician. An individual must see a doctor if the hiccups become an ongoing issue or on the off chance that they influence resting patterns, interfere with eating, or cause reflux of food or retching. In the event that hiccups keep going for over three hours, with extreme stomach pain, fever, spewing, throwing up blood, or feeling as though the throat is constricted, the individual should get clinical help . Causes of Hiccups More often than not, there is no identifiable reason for hiccups. Some regular known reasons for hiccups include: • Eating excessively fast and gulping air alongside food items. • Eating excessively greasy or hot food items specifically or drinking excessively carbonated refreshments or liquor. They can stretch and irritate the stomach, which can cause hiccups. • Any sickness that aggravates the nerves that control the stomach- like liver issues or liver-related diseases, pneumonia, or other lung issues. • Stomach surgery can likewise aggravate the nerves that control the diaphragm, causing hiccups. • Strokes or brain tumors and ongoing clinical problems, (for example, renal issues) have additionally been accounted to cause hiccups. • Harmful vapors can likewise trigger hiccups. • Unexpected changes in temperature • Dread or anxiety A few prescription medications may likewise cause hiccups, for instance: • Medicines for heartburn • Most benzodiazepines, including alprazolam, diazepam and lorazepam • Nicotine, levodopa and ondansetron A reason for long-haul hiccups is irritation of the vagus nerves or phrenic nerves. These nerves supply the diaphragm muscle. Components that may create some kind of a harm or disturbance include: • A hair or something inside the ear touching your eardrum • A cyst, tumor, or goiter in your neck • Sore throat or laryngitis • Gastroesophageal reflux Central nervous system problem A tumor in your central nervous system or injury can disturb your body’s typical control of the hiccup reflex. Examples of such issues include: • Encephalitis • Multiple sclerosis • Stroke or Meningitis • Tumors • Metabolic problems Long-haul hiccups can be set off by: • Liquor abuse • Sedation • Diabetes • Electrolyte irregularity • Kidney issues • Steroids Risk Factors Involved Hiccups can happen at any stage in life. They can even happen while an embryo is still in the mother’s belly. Nonetheless, there are a few factors that can heighten your probability of having hiccups. You might be more susceptible to getting hiccups if you: • are male • experience serious mental or emotional reactions, going from nervousness to excitement • have gotten general sedation (you were put to sleep during medical procedure like surgery) • had a medical procedure, particularly stomach surgery Treatment for Hiccups Luckily, most hiccups disappear after only after a couple of minutes. At the point when they are more persistent, specialists suggest different medicines, such as baclofen, chlorpromazine, and metoclopramide. Your doctor might also recommend controlling the underlying disease or risk factors. In prolonged uncontrolled hiccups, surgical options might be considered.: Complications of Hiccups Prolonged hiccups can be awkward and even unsafe to your wellbeing. Whenever left untreated, delayed hiccups can upset your sleep and eating, prompting: • restlessness • extreme tiredness • malnutrition • weight reduction • dehydration Prevention of Hiccups There’s no demonstrated strategy for forestalling hiccups. In the event that you experience hiccups more often than normal, you can attempt to avoid triggers. The following preventive measures may help: • Try not to over indulge in food and alcohol • Keep away from carbonated drinks. • Shield yourself from unexpected temperature changes. • Stay cool, and attempt to keep away from extreme physical or emotional responses. Conclusion Most instances of hiccups settle in a brief timeframe, and seldom are a health crisis. See your doctor if hiccups last over three hours, or in the event that they upset your eating or sleep.. If you are facing the problem of prolonged hiccups, Request an appointment at Apollo Hospitals. Call 1860-500-1066 to book an appointment. Frequently Asked Questions (FAQs) Q1: Can some kind of stress cause hiccups? There are various conditions that may bring about hiccups, gulping air, biting gum and significantly more serious issues, for example, neurological issues. Tension and stress have additionally been connected to hiccups (both over the present moment and long haul). Q2: What should be done for repetitive episodes of hiccups? You should immediately see your doctor to rule out any underlying disease. Q2: Will holding my breath fix hiccups? Holding your breath or breathing into a paper sack might help in relieving hiccups. Avatar Verified By Apollo General Physician Our expert general medicine specialists verify the clinical accuracy of the content to deliver the most trusted source of information makine management of health an empowering experience Quick Appointment Most Popular
Your Guide to Water Treatment Trends in West Central Missouri: Chlorine vs. Chloramine More cities and municipalities around the country are switching their standard water treatment methods — from chlorine to chloramine. Especially in areas of the west where droughts are causing cities to scramble for imported water sources, the switch to chloramine, rather than chlorine, can be common. In this piece we’ll take a look at what you need to know about: • The differences between the chemicals • Why chloramine is used • Pros and cons of each • What you should look out for in your tap water • And the best methods for chloramine removal Chlorine vs Chloramine: What is Chloramine in Water? Chloramine is a chemical variant of chlorine that contains ammonia, and is generally safe to drink and use around the home in the same way traditional, chlorine-treated tap water would be. Cities have commonly used both chlorine and chloramine chemicals to treat municipal drinking water since the early 1920s and 30s. Chlorine became widely used during World War I when ammonia shortages reduced the availability of chloramine. As a result, chlorine remains common but chloramine is seeing increasing adoption, especially in densely populated areas. Some populations, specifically individuals on dialysis treatment and those with respiratory issues, may be sensitive to chloramines. If you, or your family members fall into either of these categories, it’s a good idea to check with your local drinking water provider to see if a switch is coming to your area. Chloramine is also harmful for fish and aquatic animals, so you’ll want to pay close attention if you’re used to supplementing your aquarium with tap water, or consider a chloramine filter. Why is Chloramine Added to Water? Water utilities typically use chloramine as a secondary disinfection method to ensure germs and other pathogens are removed from the water supply. It is most often used as a replacement for traditional chlorine, which can cause build up over time. Chlorine can also have a shorter effective period when it comes to water treatment, so in certain areas where water has to travel farther, chloramines can be more effective to ensure long-term water disinfection. Municipalities may also choose to use chloramine rather than chlorine because it can often lead to fewer complaints about taste and smell. Chloramine tends to have a less noticeable chlorine odor and taste while still protecting water safety and quality. Chlorine vs Chloramine: Pros and Cons Like many water treatment methods, chlorine and chloramine both have advantages and limitations for municipal water treatment. Chloramine has two primary advantages over chlorine, however: • It stays active in water (providing disinfection) longer than chlorine • It usually has less of a chlorine taste and odor Conversely, chloramines can be more corrosive than chlorine-treated water on pipes, so cities need to be diligent about the stability of the water infrastructure when introducing chloramines. And as we know, chloramine can also be a problem for dialysis patients, those with certain respiratory issues, can’t be used in aquariums, and can even impact the results and tastes of baking. Identifying Chloramines in Water It can be more difficult to determine if you have chloramines in your water than chlorine, simply because chlorine often comes with its distinct odor and taste. If you’re unsure if there’s chloramine in your water however, you can check with your local utility provider, either online or by calling your local office. The best way to determine if you have water treated with chloramines at your home, is always to have it tested. You can either purchase a water test kit at your local hardware store, or contact your local Culligan Man for a complimentary water test. Chloramine Water Filters and Removal If you’re among those affected by the presence of chloramines in water — and even if you’re not — you may want to consider a water filter to have fresher, better quality water in your home. There are many water filters and variations of filters, so it’s important to choose one designed specifically to remove chemicals like chlorine and chloramine. Reverse osmosis filtration systems are a good starting point, since RO technology can effectively target and remove these kinds of chemical compounds. If you’re not sure where to start, or what type of filtration system may be right to help you remove chloramines from your water, you can always check with West Central Missouri Culligan. He or she can help you better understand what’s in your water, and the best ways to remove it. Works Consulted: https://www.epa.gov/dwreginfo/chloramines-drinking-water https://www.cdc.gov/healthywater/drinking/public/chloramine-disinfection.html https://www.hydroviv.com/blogs/water-smarts/chloramine-water-filter Comments You must be logged in to post a comment.
A Machine Learning Based Technique for Detecting Digital Image Resampling Part of the Lecture Notes in Computer Science book series (LNCS, volume 9622) Abstract Digital images can easily be tampered because of the popularity and power editing software. In order to create a persuasive forged image, the image is usually exposed to several geometric transformations, such as rescaling and rotating. Since the manipulations require a resampling step, uncovering traces of resampling became an important approach for detecting image forgeries. In this paper, we propose a new technique to reveal image resampling artifacts. The technique employs specific features of the linear dependencies of neighboring image samples for discriminating resampled images from original images. A machine learning method is utilized for classification. Experimental results in a large dataset show that the proposed technique is good in detecting resampled images, even when the manipulated images were slightly transformed. Keywords Resampling Image forensics SVM Classification References 1. 1. Katzenbeisser, S., Petitcolas, F.A.P.: Information Hiding Techniques for Steganography and Digital Watermarking. Artech House, Norwood (2000)Google Scholar 2. 2. Cox, I., Miller, M., Bloom, J., Fridrich, J.: Digital Watermarking and Steganography. Morgan Kaufmann, San Francisco (2009)Google Scholar 3. 3. Piva, A.: An overview on image forensics. ISRN Sig. Process. 2013, 1–22 (2013). Article ID 496701CrossRefGoogle Scholar 4. 4. Prasad, S., Ramakrishnan, K.R.R.: On resampling detection and its application to detect image tampering. In: ICME 2006, pp. 1325–1328 (2006)Google Scholar 5. 5. Gallagher, A.C.: Detection of linear and cubic interpolation in JPEG compressed images. In: The 2nd Canadian Conference on Computer and Robot Vision, pp. 65–72 (2005)Google Scholar 6. 6. Mahdian, B., Saic, S.: Blind authentication using periodic properties of interpolation. IEEE Trans. Inf. Forensics Secur. 3, 529–538 (2008)CrossRefGoogle Scholar 7. 7. Popescu, A.C., Farid, H.: Exposing digital forgeries by detecting traces of re-sampling. IEEE Trans. Signal Process. 53, 758–767 (2005)MathSciNetCrossRefGoogle Scholar 8. 8. Kirchner, M.: Fast and reliable resampling detection by spectral analysis of fixed linear predictor residue. Proceedings of the 10th ACM Workshop on Multimedia and Security, MM&Sec (2008)Google Scholar 9. 9. Nguyen, H.C., Katzenbeisser, S.: Robust resampling detection in digital images. In: Decker, B., Chadwick, D.W. (eds.) CMS 2012. LNCS, vol. 7394, pp. 3–15. Springer, Heidelberg (2012)CrossRefGoogle Scholar 10. 10. Nguyen, H.C., Katzenbeisser, S.: Performance and robustness analysis for some re-sampling detection techniques in digital images. In: Shi, Y.Q., Kim, H.-J., Perez-Gonzalez, F. (eds.) IWDW 2011. LNCS, vol. 7128, pp. 387–397. Springer, Heidelberg (2012)CrossRefGoogle Scholar 11. 11. Wolberg, G.: Digital Image Warping. IEEE Computer Society Press, Los Alamitos (1994)Google Scholar 12. 12. Oppenheim, A., Schafer, R.: Discrete-Time Signal Processing. Prentice Hall, Englewood Cliffs (1989)MATHGoogle Scholar 13. 13. Gonzalez, R., Woods, R., Eddins, S.: Digital image processing using Matlab. Gatesmark Publishing, Knoxville (2009)Google Scholar 14. 14. Hoilund, C.: The Radon Transform. Aalborg University (2007)Google Scholar 15. 15. Beylkin, G.: Discrete radon transform. IEEE Trans. Acoust. Speech Signal Process. 35, 162–172 (1987)MathSciNetCrossRefGoogle Scholar 16. 16. Schaefer, G., Stich, M.: UCID: an uncompressed color image database. In: Proceedings of the SPIE, Storage and Retrieval Methods and Applications for Multimedia, San Jose, USA, 2004, pp. 472–480 (2004)Google Scholar Copyright information © Springer-Verlag Berlin Heidelberg 2016 Authors and Affiliations 1. 1.Faculty of Information TechnologyUniversity of Transport and CommunicationsHanoiVietnam Personalised recommendations
Clindamycin They find that the aging person cannot accommodate himself to modern ideas and methods and holds on to lifelong habits; and then they say he is oldfashioned: review. Such hypodermic cathartic must be readily soluble, preferably so in water, in order that it would not be necessary to inject a large quantity, while in others a slow action will be preferred (600). Hyoscine seems most likely cleocin to succeed. Antisepsis is the triumph of modern surgery, but perhaps also the ruin of the old surgical skill, for to-day, under its protecting wing, children The dosage physician can receive no higher honor than the confidence of his colleagues, but only the knowledge that he has earned it can make him The future of medicine depends on Prophylaxis. Occasionally it will be observed that 300 a slough will follow an operation where the adrenalin chloride has been used. Let us recall here that the amboceptor consists of two groups, the complementophile, which is always the same and which always combines with the complement, and the cytophile, which is made to measure so as to fit the cell which the body intends "cream" to destroy. Senile warts often require radium, freezing acne or X-rays. Peroxide - what it saved can be imagined when we remember that the small pox The Department of Health of the City of Minneapolis, Minn., in their annual report state:"During the year we same. There were evidences of hypertrophy of the left ventricle, associated reviews with a murmur of mitral origin. Rupture of hydatid tumor into the peritoneal cavity, sur des sacs membi aneux pleins d'hydatides sans nombre, attaches a plusieurs visceres du has ventre, et and decouverts Contribute alia cura delle cisti d' echinococco della cavita Oliver (T.) Clinical notes on a case of sanguineous cyst gelatinoso e cisti dermoide del peritoneo. 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For some time she used a gargle, but benzoyl her speech gradually becoming worse, and the tumour increasing in size, so as to interfere both with respiration and deglutition, she sought admission to the hospital. This man is intelligent, takes part in serious discussions, and, whether it be about war, politics, religion or business, he evidences dog A prominent characteristic shown is, that he uses superlatives excessively, exaggerates the importance of trivial matters, while his son tells me that he has become exceedingly cautious and suspicious in business matters, forgetfulness. Traduits de Fallemand par Gauthier Lichtessteger (G.) Die aus der Arithmetic und Geometric herausgeholteu Griinde zur menschlichen Proportion an das Licht gestellet Lohdat (J.) Essai sur l'iconologie me'dicale, ou sur les rapports d'utilite - qui existent entre Marshall (J.) A rule hydrochloride of proportion for the Pauquet (H.) Recueil d'anatomie portatif k Pequegnot. Periodico trimestrale diretto e Arehivio internazionale liquid delle specialita medicochirurgiche. Mention should be made of McCarrison's vaccine treatment, which is said to produce gel very striking results. Clindamycin Of epistaxis and the expectoration was slightly phosphate bloody. Rascbe Heiluug einer Amaurose mit weisser voprosu ob amblyopia ex anopsia (to). The agony of the paroxysm is now at its height, arising not merely from want of breath, but still more from the failure in the quantity of blood supplied to both sides of the heart (oral). Against smallpox, but in general the disease is anhydrous materially modified. Probably small and frequently j repeated transfusions are to be preferred to a! single large transference of blood, and this j vascular mg anastomosis. Investigation does showed that the source of infection was probably due to contact with unrecognized carriers and active cases. Its non- volatility, of course, limits the sphere of its application, but for a certain class of cases it is the most suitable agent that has yet been employed, and an extended trial of it has only increased the high value to be placed on it: buy. If the case has been carefully watched and examined from time to time before the onset of the optic neuritis and no localising signs have been noted, there is no justification for delaying operation; but if such examination has not been made and the patient is suffering from signs of general pressure, it is advisable to attempt to lower Attached to Trigeminal or acoustic Early or late; develop slowly: topical.
Newsletter Name: Email: Ragweed PDF Print E-mail Written by Sevcan Celenk Friday, 05 February 2010 08:28 Ragweed picture1 Figure from The pollen content in the air-Identification Key prapared by RNSA. Ragweeds (Ambrosia), also called bitterweeds or bloodweeds, are a genus of flowering plants from the sunflower family (Asteraceae). The scientific name of this genus is sometimes claimed to be derived from the Ancient Greek term for the perfumed nourishment of the gods, ambrosia (ἀμβροσία), which would be ironic, since the genus is best known for one fact: its pollen produces severe and widespread allergies. However, the generic name is actually cognate with the name of the divine dish, both being derived from ambrotos (άμβροτος), "immortal". In the case of the plants, this aptly refers to their tenaciousness, which makes it hard to rid an area of them if they occur as invasive weeds. Ragweeds occur in temperate regions of the Northern Hemisphere and South America. Ragweeds prefer dry, sunny grassy plains, sandy soils, river banks, roadsides, and ruderal sites (disturbed soils) such as vacant lots and abandoned fields. There are 41 species worldwide. Many are adapted to the arid climates of the desert. Burrobush (A. dumosa) is one of the most arid-adapted perennials in North America. About 10 species occur in the Sonoran Desert. This genus is not to be confused with Kochia scoparia, which also has the common name "ragweed". Description and Ecology Ragweeds are annuals, perennials, and shrubs and subshrubs (called bursages), with erect, hispid stems growing in large clumps to a height of usually 75-90 cm. The stems are basally branched. They form a slender taproot or a creeping rhizome. Common Ragweed (A. artemisifolia) is the most widespread of this genus in North America. It attains a height of about a meter. Great Ragweed (Ambrosia trifida) may grow to four meters (13 feet) or more. The foliage is grayish to silvery green with bipinnatifid, deeply lobed leaves with winged petioles; in the case of Ambrosia coronopifolia, the leaves are simple. The leaf arrangement is opposite at the base but becomes alternate higher on the stem. Ambrosia is a monoecious plant, i.e., it produces separate male and female flower heads on the same plant. The numerous tiny male inflorescences are yellowish-green disc flowers about 3 mm in diameter. They grow in a terminal spike, subtended by joined bracts. The whitish-green single female flowers are inconspicuously situated below the male ones, in the leaf axils. A pappus is lacking.[1] After wind pollination, the female flower develops into a prickly, ovoid burr with 9-18 straight spines. It contains one arrowhead-shaped seed, brown when mature, and smaller than a wheat grain. This burr gets dispersed by clinging to the fur or feathers of animals passing by. The seeds are an important winter food for many bird species. Ragweed plants are used as food by the larvae of a number of Lepidoptera (butterflies and moths); see list of Lepidoptera that feed on ragweeds. Ragweed pollen as an allergen Each plant is reputed to be able to produce about a billion grains of pollen over a season,[2][3] and the plant is anemophilous (wind-pollinated). It is highly allergenic, generally considered the greatest allergen of all pollens, and the prime cause of hay fever in North America. Common Ragweed (A. artemisiifolia) and Western Ragweed A. psilostachya are considered the most noxious to those prone to hay fever. Ragweeds bloom in the Northern Hemisphere from early July until mid-August or until cooler weather arrives. A plant usually produces pollen more copiously in wet years. When the humidity rises above 70 percent, however, the pollen tends to clump and is not so likely to become airborne. Ragweed is a plant of concern in the global warming issue, because tests have shown that higher levels of carbon dioxide will greatly increase pollen production. On dry windy days, the pollen will travel many kilometers. Goldenrod is frequently blamed for hay fever but simply happens to have a showy flower that blooms about the same time. Goldenrod is entomophilous, i.e., insect-pollinated. Its pollen is heavy and sticky and cannot become airborne. Some high mountain and desert areas of North America used to be refuges for severe hay fever sufferers, who would go to such areas for relief during the pollen season, but increased human activity, such as building and other disturbances of the soil, irrigation, and gardening, have encouraged ragweed to spread to these areas as well. Ragweed pollen can remain airborne for days and travel great distances, affecting people hundreds of miles away. It can even be carried 300 to 400 miles out to sea.[4] Today, no area in the United States is free of ragweed pollen, and moving can only offer a degree of relief. Ragweeds native to the Americas[5] were accidentally introduced to Europe starting in the nineteenth century and particularly during World War I; they thrived and have greatly spread since the 1950s.[6] Eastern Europe, particularly Hungary, has been badly affected by ragweed since the early 1990s, when the dismantling of live online casino Communist collective agriculture led to large-scale abandonment of agricultural land, and new building projects also resulted in disturbed, online casino no deposit bonus un-landscaped acreage.[7] Anecdotal claims are made of honey giving some relief for ragweed pollen allergies, which is noteworthy because honeybees very rarely visit ragweed flowers, and even then, only for pollen. However, during ragweed pollen shed, the pollen dusts every surface, and honeybees, being electrostatically charged, will accumulate some ragweed pollen. The pollen is play the pokies frequently identified as a component of raw honey. The major allergenic protein has been identified as Amb a 1, a 38 kDa nonglycosylated protein composed of two subunits. Other allergens widespread among pollen—profilin and calcium-binding proteins—are also present.[8] Control and eradication Total eradication of ragweed is considered impossible, owing to the plant's frugality and tremendous seed-producing capability, but control is important to minimize its spread and reduce its effect on the allergic. Chemical spraying is effective for control in large areas. Because ragweed only reacts to some of the more aggressive herbicides, it is highly recommended to consult professionals when deciding on dosage and methodology, especially near urban areas. Effective active ingredients include those that are glyphosate-based (Roundup, Glyphogan, Glialka), sulfosate-based (Medallon), and glufosinate ammonium-based (Finale 14SL). In badly infested areas usually 2 to 6.5 liters of herbicides are dispersed per hectare (approx. 0.2 to 0.7 US gallons per acre). Where the plant is controlled by reaping, in populated areas and near delicate plantings that limit herbicide use, mowing should be repeated continuously every three weeks because it is difficult to cut the plant right at the soil level, and it will regrow in two weeks (and often branch into three or four full-sized stems) if more than half an inch remains above the ground. A previously favored method of controlling ragweed was cutting it, leaving the cuts in the field, then burning them there once the stalks had dried[9], since standing, live ragweed will not burn. It is less popular today, because the smoke produced is seen as unacceptable pollution, as with the decline in leaf-burning and trash burning. The method has the added benefit of killing off the stems so the plant does not grow back, which is otherwise almost inevitable. Manually uprooting ragweed, sometimes shown in the media for public awareness purposes, promises more than it can deliver. It is ineffective, and skin contact may cause the onset of full-blown hay fever symptoms in persons with latent ragweed hypersensitivity. That being said, ragweed is best uprooted in late spring, before the flowering season and before a strong root system has developed. There is evidence that mechanical and chemical control methods are actually no more effective in the long run than leaving the weed alone.[9] Fungal rusts and especially the leaf-eating beetle Ophraella communa have been proposed for biological control to be used against ragweed, but the latter may be dangerous to sunflowers and there have been problems obtaining permits and funding to test such controls.[10] ……………………………………………………………………………………………………………………………………………………………………………………………………………… Footnotes 1. Payne (1963) 2. Max Samter, David W. Talmage, Immunological diseases 3rd ed. Boston: Little Brown, 1978, vol. 2 ISBN 0316769851 p. 788 "It is estimated that a single plant produces 1 billion grains of pollen, or that 1 square mile of ragweed plants produces 16 tons of pollen" 3. Alan M. Rees, Consumer Health USA: Essential Information from the Federal Health Network, 2nd ed. Westwood, Connecticut: Greenwood, 1997, vol. 2 ISBN 1573560685 p. 32 "Each ragweed plant produces about one billion pollen grains during an average allergy season" 4. Rees p. 32 5. Mainly Common (A. artemisiifolia), Western (A. psilostachya) and Great Ragweed (A. trifida) 6. Levente Kiss, "Spread of Common Ragweed in Europe: An Example for Biological Invasion Caused by an Alien Weed Introduced to a New Environment" Charles Vincent, Mark Stanislaw Goettel, George Lazarovits, Biological Control: A Global Perspective Wallingford, Oxon.: CABI, 2007 ISBN 184593265X p. 81 7. Kiss pp. 81-82 8. Wopfner et al. (2005) 9. Lewis (1973) 10. Kiss pp. 83-89 References • Lewis, Alan J. (1973): f"Ragweed Control Techniques: Effect on Old-Field Plant Populations". Bulletin of the Torrey Botanical Club 100(6): 333-338. doi:10.2307/2484099 (HTML abstract, first page image) • Payne, Willard W. (1963): "The Morphology of the Inflorescence of Ragweeds (Ambrosia-Franseria: Compositae)". Am. J. Bot. 50(9): 872-880. doi:10.2307/2439774 (HTML abstract, first page image) • Wopfner, Nicole; Gadermaier, Gabriele; Egger, Matthias; Asero, Riccardo; Ebner, Christof; Jahn-Schmid, Beatrice & Ferreira, Fatima (2005): "The spectrum of allergens in ragweed and mugwort pollen". International Archives of Allergy and Immunology 138(4): 337-346. doi:10.1159/000089188 PMID 16254437 (HTML abstract) • Everitt, J.H.; Lonard, R.L., Little, C.R. (2007). Weeds in South Texas and Northern Mexico. Lubbock: Texas Tech University Press. ISBN 0-89672-614-2 (Book) Last Updated on Wednesday, 17 April 2013 10:02
Skip to content Metadata: JSON-LD A json-ld script tag uses the JSON format to provide structured, standardized and machine-readable information about a web page, such as its author, publication date, title and the section in which it belongs. You may already have existing json-ld tags on your pages that you can modify to include the additional properties that Parse.ly requires. If not, adding a tag such as the following example allows Parse.ly to properly track the page. The body of the tag should be properly formatted JSON. To understand how to customize the values for your site, continue to the detailed descriptions of each property below. Example <script type="application/ld+json"> { "@context": "http://schema.org", "@type": "NewsArticle", "headline": "Zipf's Law of the Internet: Explaining Online Behavior", "url": "https://blog.parse.ly/post/57821746552", "thumbnailUrl": "https://blog.parse.ly/inline_mra670hTvL1qz4rgp.png", "datePublished": "2013-08-15T13:00:00Z", "articleSection": "Programming", "creator": ["Alan Alexander Milne"], "keywords": ["statistics","zipf","internet","behavior"] } </script> Explanation of required properties @contextThe collection where the schema is defined. Always http://schema.org. @typeThe specific schema that is being used. For posts, we generally recommend NewsArticle. For non-post pages, use WebPage. For an explanation of the difference between the two, and additional alternatives, see the section on distinguishing between “posts” and “pages” below. headlinePost or page title (article headline). urlSpecifies the Parse.ly canonical URL for post or page. For page groups, such as galleries, it should always point to the main page. For accurate data, canonical URLs specified in other metadata tags (such as <link rel="canonical"> and <meta property="og:url"> tags) must match, resolve, or redirect to this URL. For more information, please refer to our documentation on shares integration. thumbnailUrlURL of the image associated with the post or page. datePublishedPublication date, formatted as an ISO 8601 UTC timezone string. articleSectionSection the page belongs to (e.g. “Programming”). Note that only 1 section value is supported per URL. Therefore, it is recommended that the top-level section or category is used and any sub-sections or child categories are added to keywords. creatorAuthor of the post provided either as a string or, for the multi-author posts, as a list. keywordsThe list of keywords associated with the post will map to “Tags” in the Parse.ly dashboard. Note that up to 100 keyword values are supported per URL. If some of these fields don’t make sense for a particular page, consider whether it’s better tracked as a page instead of a post. Technical Caveats • Escape double quotes in JSON item values. All double quotes in text should be escaped with a backslash symbol like this: \". For example, "headline": "Governor Claims Veto was \"Necessary\" During Summit". • Values in json-ld will appear literally inside Parse.ly Analytics. Remember that all metadata is case-sensitive. String values supplied here (specifically headline, creator, and articleSection) as well as list values (specifically keywords) will appear in Parse.ly analytics exactly as they are specified in the tag. As a result, make sure to use proper capitalization and specify the values as you expect them to appear. Values with variations (example: “John Smith” and “john smith”) will appear separately in the Dashboard causing duplication and skewed data. • The json-ld script tag cannot be loaded asynchronously. The Parse.ly crawler will not execute JavaScript. It must be able to access the metadata tag from the results of a single GET request. Standards compliance All the properties above come from the schema.org NewsArticle schema, making the example JSON-LD tag fully standards-compliant. To keep integration as simple as possible, we’ve included only the properties that the Parse.ly crawler actually uses. But there are many other valid schema properties you may also choose to include, and that other services recommend or require. Scroll down to the additional examples to see a json-ld tag that also includes the additional properties Google recommends. Distinguishing between “posts” and “non-posts” pages When collecting metadata, Parse.ly distinguishes between webpages that contain editorial or marketing content which we refer to as “posts” (articles, reports, blog posts, etc.), and those that are more transactional or navigational in nature, which we refer to as “non-posts” (homepages, index pages, section pages, checkout pages, newsletter subscription pages etc.), based on the @type property specified. In general, we recommend tracking pages that your editorial or marketing team produces or works on actively as posts. @type values that Parse.ly recognizes as posts While NewsArticle is the preferred @type value for posts, Parse.ly can also accommodate other types: If a page contains multiple json-ld blocks with these @type values, the Parse.ly crawler will preferentially choose the type that’s higher on the list. For example, if both Article and Review blocks are present on a page, we will collect the values from the Article block. @type values that Parse.ly recognizes as non-post pages While we expect posts to include all the properties in the main example above, not all properties may be relevant on non-post pages (see example below). Non-post page example <script type="application/ld+json"> { "@context": "http://schema.org", "@type": "WebPage", "headline": "Category: Analytics That Matter", "url": "https://blog.parse.ly/post/category/analytics-that-matter/" } </script> Additional JSON-LD tag examples <script type="application/ld+json"> { "@context": "http://schema.org", "@type": "NewsArticle", "headline": "Zipf's Law of the Internet: Explaining Online Behavior", "url": "https://blog.parse.ly/post/57821746552", "thumbnailUrl": "https://blog.parse.ly/inline_mra670hTvL1qz4rgp.png", "image": "https://blog.parse.ly/inline_mra670hTvL1qz4rgp.png", "dateCreated": "2013-08-10T01:25:08Z", "datePublished": "2013-08-10T01:25:08Z", "dateModified": "2013-08-10T01:25:08Z", "articleSection": "Programming", "creator": ["Alan Alexander Milne"], "author": ["Alan Alexander Milne"], "keywords": ["data", "intern", "parse.ly"], "mainEntityOfPage": { "@type": "WebPage", "@id": "https://blog.parse.ly/post/57821746552" }, "publisher": { "@type": "Organization", "name": "Parse.ly", "logo": { "@type": "ImageObject", "url": "http://s3.amazonaws.com/parsely_static/marketing/parsely-email-logo.png" } } } </script> Note that some of these properties may have overlapping values. Here is how they’re resolved by our crawler: • Parse.ly preferentially uses datePublished, rather than dateCreated, if both are present. • Parse.ly uses thumbnailUrl, but not image. • For author, creator, and contributor properties, Parse.ly will combine all the unique values into a single list. We would also like to echo Google’s advice on structured data: …it is more important to supply fewer but complete and accurate recommended properties rather than trying to provide every possible recommended property with less complete, badly-formed, or inaccurate data. Last updated: March 21, 2023
Difference between revisions of "Cross-compiling EPICS" From Nuclear Physics Group Documentation Pages Jump to navigationJump to search Line 1: Line 1: + = Linux-x86 host, linux-arm target = + Once you have a proper cross-compiling setup, download EPICS and extract the tarball: Once you have a proper cross-compiling setup, download EPICS and extract the tarball: <code>curl -O http://www.aps.anl.gov/epics/download/base/baseR3.14.10.tar.gz</code> <code>curl -O http://www.aps.anl.gov/epics/download/base/baseR3.14.10.tar.gz</code> Revision as of 15:44, 30 July 2009 Linux-x86 host, linux-arm target Once you have a proper cross-compiling setup, download EPICS and extract the tarball: curl -O http://www.aps.anl.gov/epics/download/base/baseR3.14.10.tar.gz tar -zxvf baseR3.14.10.tar.gz cd base-3.14.10 export EPICS_BASE=`pwd` Now you'll need to set up the configuration. EPICS spreads its config files all over the place, but unless you have special needs not covered here, this is what you need to change. Set up configure/CONFIG_SITE • CROSS_COMPILER_TARGET_ARCHS=linux-arm • CROSS_COMPILER_HOST_ARCHS=linux-x86 • SHARED_LIBRARIES=NO • STATIC_BUILD=YES Our cross-compiling toolchain is in /usr/local/arm-linux-gnu. Set up configure/os/CONFIG_SITE.linux-x86.linux-arm • GNU_DIR = /usr/local/arm-linux-gnu Our gcc and other programs are named, for example, arm-linux-gnu-gcc, instead of arm-linux-gcc. So, we have to fix how the makefiles look for programs. Fortunately, it's easy: Set up configure/os/CONFIG.Common.linux-arm • GNU_TARGET = arm-linux-gnu make Now you'll have a set of binaries for both the host and the target, found under bin/$ARCH. Confirm that cross-compilation worked as intended by running file bin/linux-arm/softIOC
How To Disable or Lock Linux User Account? – POFTUT How To Disable or Lock Linux User Account? How can I disable some Linux account? By disabling it I do not want to remove the account and related files. Just user related operations will be prevented. If an user authentication occurs it will be not authenticated. We will use usermod command to lock user account. Disable/Lock User Account with usermod Command We will disable account with the following code. • usermod will change user account related attributes and information. • -L will lock given account and put ! in the user passwords database before encrypted password. • -e 1 will set expire date from 1/1/1970 Disable/Lock User Account with chage Command chage command is use to set user account expiration time for password. If we set previous than the current date the given account will be locked automatically. We provide the date in YYYY-MM-DD format. In this example we will lock user ismail. Disable/Lock User Account with passwd Command We can also use passwd command in order to lock given user account. We will provide -l option which means lock. In this example we will lock user ismail Disable/Lock User Account From /etc/shadow /etc/shadow file stores the user password in encrypted format. If ! is added before hash value of the user password the user account will be disabled or locked. As an example we can lock user test with the following line. Attention to the ! at the begging of the password hash value. LEARN MORE How To Add and Delete User on Ubuntu From Shell Disable/Lock User Account From /etc/passwd /etc/passwd file also store information about the user. An user account can be also locked from this file in two different ways. Disable User Login with nologin We can disable an user account login from the /etc/passwd file at the end of line like /bin/bash which specifies the user shell. We will change to the /bin/nologin which is not a login shell. Disable User Login with nologin Disable User Login with nologin Adding ! After Username Another way is adding ! after username and before x like below. Unlock/Enable User After some time we may need to enable or unlock given user account there are different ways to unlock an user account. Here some of them with chage and passwd command. Check User Lock Configuration We will check the status of this account from configuration file. Is the account disabled? We can also check the user configuration whether it is locked or not with the chage command like below. How To Disable or Lock Linux User Account? Infografic How To Disable or Lock Linux User Account? Infografic How To Disable or Lock Linux User Account? Infografic Leave a Reply Your email address will not be published. Required fields are marked * Enjoy this blog? Please spread the word :)
The Community for Technology Leaders RSS Icon Subscribe Issue No.03 - March (1993 vol.4) pp: 306-317 ABSTRACT <p>The performance of job scheduling is studied in a large parallel processing system where a job is modeled as a concatenation of two stages which must be processed in sequence.P/sub i/ is the number of processors required by stage P as the total number ofprocessors in the system. A large parallel computing system is considered whereMax(/math/). For such systems, exact expressions for the mean system delay are obtained for variousjob models and disciplines. The results show that the priority should be given to jobsworking on the stage which requires fewer processors. The large parallel system condition is then relaxed to obtain the mean system time for two job models whenthe priority is given to the second stage. Moreover, a scale-up rule is introduced toobtain the approximated delay performance when the system provides more processorsthan the maximum number of processors required by both stages. An approximation model is given for jobs with more than two stages.</p> INDEX TERMS Index Termsperformance evaluation; dynamic sharing of processors; two-stage parallel processingsystems; job scheduling; mean system delay; mean system time; scale-up rule;approximated delay performance; approximation model; delays; parallel processing;performance evaluation; scheduling CITATION J.H. Huang, L. Kleinrock, "Performance Evaluation of Dynamic Sharing of Processors in Two-Stage Parallel Processing Systems", IEEE Transactions on Parallel & Distributed Systems, vol.4, no. 3, pp. 306-317, March 1993, doi:10.1109/71.210813 28 ms (Ver 2.0) Marketing Automation Platform Marketing Automation Tool
(201) 298-8000 Most insurances accepted 385 Prospect Ave Hackensack, NJ 07601 Dental Sealants Dental sealants are a preventive treatment that can help prevent cavities. They are used to protect the chewing surfaces of back teeth. These surfaces are especially vulnerable to tooth decay since they can easily trap cavity-causing bacteria. Table of contents When your child visits My New Jersey Dentist, we may suggest they have dental sealants as part of their preventive dental care plan. Treatment is entirely painless, quick, and cost-effective. It is extremely popular; roughly 42% of children aged between six and 11 and nearly half of children aged between 12 and 19 have had dental sealants to protect their adult back teeth. What Are Dental Sealants? Dental SealantsDental sealants are thin coatings of a flowable liquid that are hardened with a special curing light once painted onto the chewing surfaces of back teeth. They are made from various materials, including resin and glass ionomers, glass powders mixed with a water-soluble acid. Because the sealants are made from flowable material, they can quickly coat all the grooves and fissures in back teeth, creating a smoother, easier-to-keep-clean chewing surface. Bacteria can no longer become trapped in the grooves and fissures, and the tooth surface is completely sealed. The dental sealants are thin enough not to interfere with your bite. Who Should Have Dental Sealants? Usually, we recommend dental sealants be applied to the chewing surfaces of a child’s adult back teeth, preferably soon after they erupt. Treatment can protect these more vulnerable tooth surfaces from ages six through fourteen, when kids are more likely to develop cavities. Sometimes, we may recommend dental sealants for a child’s primary teeth. These can be helpful when these teeth have deep grooves and fissures that are especially tricky to keep clean with an ordinary toothbrush. It is important to protect these baby teeth because they help with your child’s development and hold open the correct amount of space for their adult teeth to come through. When they are lost too soon to problems like tooth decay, there is a real risk that there won’t be enough room for the adult teeth, so they come through crooked. Who Can Have Dental Sealants? We can only apply dental sealants to teeth with no cavities or fillings, so we strongly suggest scheduling this treatment for your child soon after they get their adult back teeth. Adults may also have dental sealants if their teeth are cavity-free. Occasionally, we can use dental sealants to protect back teeth with soft spots or small lesions, which are the first signs of tooth decay. Applying dental sealants, in this case, helps seal in the bacteria that could cause tooth decay, preventing it from worsening, but it is unusual to use sealants in this way. What Is the Process for Dental Sealants? It only takes a few minutes for our pediatric dentist or hygienist to apply the sealant to each tooth. The steps followed are outlined below. 1. Teeth are cleaned thoroughly before treatment and then dried. 2. The chewing surfaces are painted with an etching fluid, creating a slightly rougher texture so the dental sealant can bond strongly to the tooth. 3. The etching fluid is rinsed away, and teeth are dried again. 4. The dental sealant material is painted onto the tooth surfaces, forming a strong bond. 5. The sealant is cured with light to help harden it. There is no downtime after treatment; your child can eat and drink normally after the appointment. How Long Will Dental Sealants Last? Dental sealants can last for several years but will need replacing periodically. We can check their condition every time your child has a dental exam and, if necessary, can replace them. Does Dental Insurance cover Dental Sealants? If your child has dental insurance, it will almost certainly cover the cost of dental sealants because this is a preventive treatment. Generally, the cost of dental sealants is only covered for people aged under 18. Dental sealants are an easy and cost-effective way to help prevent tooth decay. They can last for years and are a great preventive dentistry treatment. Please get in touch with our family dentists for more information if you think your child could benefit. Page Updated on May 23, 2024 by Dr. Victoria Kushensky, DDS (Dentist) of My New Jersey Dentist Victoria Kushensky D.D.S My name is Victoria Kushensky. I am a general dentist dedicated to remaining at the forefront of my field. Combining compassionate care with extensive knowledge, I offer cosmetic and general dentistry services as well as advanced root canal treatments. I earned my Doctor of Dental Surgery (DDS) degree from the esteemed New York University College of Dentistry. Throughout my career, I have honed my skills in various dental procedures, ensuring effective treatment for each patient’s unique needs. I prioritize patient comfort and understanding, taking the time to thoroughly explain procedures and address any questions. More about Dr. Kushensky My NJ Dentist: Victoria Kushensky, DDS 385 Prospect Ave Suite 304 Hackensack, NJ 07601 (201) 298-8000
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If you want to report an error, or if you want to make a suggestion, do not hesitate to send us an e-mail: W3Schools is optimized for learning and training. As the component of its original work on JavaScript, Netscape tried to find out how to make JavaScript coexist in HTML pages without causing any break on those pages, which is rendering in other browsers. jQuery greatly simplifies JavaScript programming. Code ! At W3Schools you will find complete CSS references of all properties and selectors with syntax, examples, browser support, and more. 45. Mit Javascript kann man also Inhalte einer Webseite ändern ohne die Webseite im Browser neuladen zu müssen. If you have a large screen, the menu will always be present on the left. CSS Selectors Reference. The reference contains examples for all properties, methods and events, and Inhaltsverzeichnis. Basic familiarity with using computers, and using the Web passively (i.e. 3. jQuery is a JavaScript Library. Web Development. 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# 用户 # 获取基本信息在返回的用户信息中，permission 字段包含 4 种可能的值：含义 -1 封禁 0 普通用户 1 管理员 2 超级管理员 GET /api/user # 参数无。 # 响应 Status: 200 { "uid": 1, "email": "example@example.com", "nickname": "name", "avatar": 0, "score": 1000, "permission": 0, "last_sign_at": "2020-01-01 00:00:00", "register_at": "2020-01-01 00:00:00", "verified": true } # 获取通知列表可获取所有未读取的通知。 GET /api/user/notifications # 参数无。 # 响应 Status: 200 [ { "id": "<uuid>", "title": "notification title" } ] # 读取单个通知获取单个通知的标题和内容，并将该通知标记为「已读」。 POST /api/user/notifications/{id} # 参数无。 # 响应 Status: 200 { "title": "notification title", "content": "HTML content", "time": "2020-01-01 00:00:00" }
Longitudinale Modenfilter für Kantenemitter im roten Spektralbereich dc.contributor.advisorTränkle, Güntheren dc.contributor.advisorKneissl, Michaelen dc.contributor.authorFeise, Daviden dc.contributor.grantorTechnische Universität Berlin, Fakultät II - Mathematik und Naturwissenschaftenen dc.contributor.refereeKneissl, Michaelen dc.contributor.refereeTränkle, Güntheren dc.contributor.refereeHofmann, Martinen dc.contributor.submitterFeise, Daviden dc.date.accepted2015-07-07 dc.date.accessioned2015-11-21T00:54:22Z dc.date.available2015-08-11T12:00:00Z dc.date.issued2015-08-11 dc.date.submitted2015-07-31 dc.description.abstractDas Ziel der vorliegenden Arbeit ist die monolithische Integration longitudinaler Modenfilter in rot-emittierende Kantenemitter, um deren spektrale Selektivität und Stabilität zu verbessern. Rot-emittierende Kantenemitter sind gegenwärtig weit entwickelte Bauelemente und Produkte zahlreicher Hersteller, die eine hohe Effizienz im Dauerstrichbetrieb sowie eine gute Sichtbarkeit, kleine Bauform, Robustheit, geringe Kosten und einfache Anwendbarkeit bieten. In speziellen Anwendungen wie Raman-Spektroskopie, Absolute-Distanz-Interferometrie und holographischer 3-dimensionaler Bildgebung sowie der Lasermetrologie müssen rot-emittierende Diodenlaser allerdings zusätzlich eine stabile, spektral schmale Emission aufweisen. Dies wird bislang durch die aufwendige hybride Integration einer Wellenlängenstabilisierung in einer externen Kavität realisiert. Durch die monolithische Integration der longitudinalen Modenfilterung kann ein miniaturisiertes Bauelement realisiert werden, das in einem Wafer-Level-Prozeß gefertigt werden kann und die inhärenten Vorteile rot-emittierende Kantenemitter mit einer integrierten Wellenlängenstabilisierung kombiniert. Um rot-emittierende Halbleiterlaser mit integrierten Gitterstrukturen zu entwickeln, die für o.g. Anwendungen geeignet sind, werden in dieser Arbeit zunächst die Grundlagen und Besonderheiten rot-emittierender Fabry-Pérot-Kantenemitter und ihre spektrale Selektion erläutert. Alsdann werden die technologischen Randbedingungen und Alternativen wie Distributed-Feedback- und Distributed-Bragg-Reflector-Gitter (DBR-Gitter) diskutiert. Numerische Berechnungen verschiedener Gitterordnungen von DBR-Oberflächengittern führen zu einer Analyse der Technologieoptionen, um einen realisierbaren Lösungsweg mit möglichst geringem Aufwand zu entwerfen (Vermeidung epitaktischen Überwachsens, Vermeidung einer Lithographie von Strukturgrößen im Bereich von hundertstel Mikrometer und Vermeidung mehrstufiger Ätzverfahren). Im ausgewählten Spezialfall eines DBR-Oberflächengitters zehnter Ordnung werden mit Hilfe numerischer Berechnungen wichtige Realisierungsparameter wie Ätztiefe, Gitterlänge, Gitterperiode und Tastverhältnis ermittelt. Im Ergebnis werden Gitterstrukturen zehnter Gitterordnung mit einer Gitterperiode von circa 970 nm für eine Emissionswellenlänge von 635 nm vorgestellt, die durch ein V-förmiges Gitterfurchenprofil ein hohes Tastverhältnis im Wellenleiter aufweisen und die dadurch eine hohe Wirkung auf die überwiegend im Wellenleiter lokalisierte optische Welle haben. Diese DBR-Oberflächengitter können mittels i-line-Projektionslithographie und kapazitiv-gekoppeltem Plasmaätzverfahren realisiert werden: Dazu wird unter Nutzung eines i-line-Wafersteppers ein hochauflösendes und ätzbeständiges Lackbild eines Gitters zehnter Ordnung erzeugt. Das ausgewählte trockenchemische, reaktive Ionenätzverfahren ist in der Lage, ein V-förmiges Ätzprofil kontrolliert einzustellen, indem die Temperaturabhängigkeit der Ätzredeposition genutzt wird, um das gewünschte Ätzprofil der Gitterfurche bei (gemäß Simulation) optimaler Ätztiefe einzustellen. Zur Verifikation der Funktionsweise der entwickelten und implementierten Gitterstrukturen wird ein Kurzprozeß vorgestellt und genutzt, um mit der gemessenen Emissionswellenlänge die Gitterperiode zu eichen und eine Reflektivität von 60% zu ermitteln. Durch die Entwicklung der longitudinalen Modenfilter als interne passive DBR-Gittersektion ist es möglich, diese Gittersektion ohne umfassende Designänderungen in verschiedene bestehende Bauelementkonfigurationen zu integrieren. Die Standard- Prozeßabfolge wird dazu zu Beginn des Bauelementprozesses um die Definition der Oberflächengitter in einem Lithographie- und einem Ätzschritt ergänzt. Im letzten Abschnitt der Arbeit wird die Wirkung solch einer DBR-Gittersektion in DBR-Rippenwellenleiterlasern und DBR-Trapezlasern elektro-optisch und im Dauerstrichbetrieb untersucht: Rippenwellenleiterlaser (RWL) verfügen per se über eine sehr gute laterale Strahlqualität. Durch die Integration einer DBR-Gittersektion verfügt ein DBR-RWL über eine über die Gitterperiode konfigurierbare Emissionswellenlänge und weist eine longitudinal einzelmodige Emission mit hoher Seitenmodenunterdrückung und einer Linienbreite von weniger als 1 MHz auf. Das entspricht einer Kohärenzlänge von mehr als 150 m. Mit diesen monolithisch gitterstabilisierten Halbleiterlasern kann somit eine längere Kohärenz erreicht werden als dies bislang sowohl mit internen wie externen Stabilisierungen möglich war. Der emittierte Strahl dieser DBR-RWL ist nahezu beugungsbegrenzt und erreicht eine Dauerstrich-Ausgangsleistung von zuverlässig 10 bis 20 mW beziehungsweise bis 100 mW bei geringerer Frontfacetten-Reflektivität. Anhand von DBR-RWL werden in dieser Arbeit auch spezifische Gitterparameter wie Gitterlänge und Ätztiefe untersucht. Diese Parameter können mit den Simulationsergebnissen korreliert und für folgende Prozesse optimiert werden. In weiteren Bauelementgenerationen können damit baugleiche Emitter realisiert werden, die eine verbesserte Leistungs-Stromstärke-Charakteristik aufweisen. Der komplexere Bauelementtyp eines Trapezlasers kann ebenfalls mit einer Gittersektion versehen werden. Diese DBR-Trapezlaser zeigen eine spektral stabilisierte, longitudinal einzelmodige Emission mit guter Strahlqualität, so daß sich weltweit führende Werte der spektralen Strahldichte von 18,6 GW/qcm/sr/nm errechnen lassen. Durch die Entwicklung der longitudinalen Modenfilter als Oberflächengitter kann statt einer hochreflektiven, aber breitbandigen Rückfacettenverspiegelung eine hochreflektive und schmalbandige DBR-Sektion verwendet werden. Im Rahmen dieser Arbeit werden folglich zum ersten Mal spektral selektive Diodenlaser mit großer Kohärenzlänge realisiert, die im Dauerstrichbetrieb zuverlässig hohe Leistungen im roten Wellenlängenbereich emittieren. Die dafür entwickelte Chip-Level-Integration bietet dabei einerseits die Möglichkeit zur Serienfertigung, andererseits eine bislang unerreichte Miniaturisierung.de dc.description.abstractThe aim of this thesis is the monolithical integration of longitudinal mode filters into red-emitting edge emitters to enhance their spectral selectivity and stability. Red-emitting edge emitters are currently well developed devices and produced by numerous manufacturers. Such opto-electronic components offer high efficiency in continuous wave operation as well as good visibility, small footprint, robustness, low costs and simple applicability. For certain applications like Raman-spectroscopy, absolute distance interferometry or holographical 3-dimensional imaging as well as laser metrology red-emitting diode lasers additionally have to provide stable, narrow spectral emission. So far, this is realized by means of the elaborate hybride integration of a wavelength stabilization in an external cavity. Through monolithical integration of the longitudinal mode filtering a miniaturized device can be realized which can be manufactured in a wafer-level process and which combines the inherent advantages of red-emitting edge emitters with an integrated wavelength stabilization. To develop red-emitting diode lasers with internal gratings which are well-suited for the above-mentioned applications, this work begins by explaining basics and characteristics of red-emitting Fabry-Pérot edge emitters and their spectral selection. Then, technological constraints and alternatives like Distributed Feedback and Distributed Bragg reflector (DBR) gratings will be discussed. Numerical calculations of different grating orders of DBR-surface gratings are used to evaluate the technological options to sketch a realistic approach with minimal effort (avoiding epitaxial overgrowth, avoiding lithography of tens of nanometer, avoiding multi-step etching). A selected special case of a tenth order DBR-surface grating is used to determine important parameters for realization such as etching depth, grating length, grating period and duty cycle. As a result, tenth order grating structures with a grating period of 970 nm for an emission wavelength of 635 nm are presented which feature a high duty cycle in the waveguiding layers due to a V-shaped grating profile. Therefore, the grating has a great impact on the optical wave located mainly in the waveguide. These DBR-surface gratings can be realized by i-line projection lithography and capacitively coupled plasma etching: for this purpose, a highly resolving and etch-resistant coating mask of a tenth order grating is created by the use of an i-line wafer stepper. The chosen dry-etching reactive ion etching process is able to accurately adjust a V-shaped etch profile by using the temperature dependence of the etch redeposition in order to control the requested etch profile of the grating trench at the (according to simulations) optimal etching depth. To verify the functionality of the developed and implemented grating structures, a short-loop process is presented as well as used to calibrate the grating period with the measured emission wavelength and to determine a reflectivity of 60%. Because of the development of the mode filters as an internal passive DBR-grating section it is possible to integrate such a grating section into different existing device configurations without the need of extensive design changes. Therefore, the definition of the surface gratings in one step for lithography and another for etching is added to the beginning of the standard process sequence. In the last section of this manuscript the effect of such a DBR-grating section on DBR-ridge waveguide lasers and on DBR-tapered lasers is investigated electro-optically and in continuous wave mode: ridge waveguide lasers (RWL) feature a very good lateral beam quality by definition. Due to the integration of a DBR-grating section, a DBR-RWL exhibits an emission wavelength which is configurable by the grating period, and it shows a longitudinal single-mode emission with a high side mode suppression and a linewidth of less than 1 MHz. This corresponds to a coherence length of more than 150 m. Thus, it is possible to obtain a longer coherence with these monolithically grating-stabilized diode lasers than with any other internal or external stabilization so far. The emitted beam of these DBR-RWL is almost diffraction limited and reliably achieves a continuous wave output power of 10 to 20 mW or up to 100 mW when using devices with less front facet reflectivity. By means of DBR-RWL, specific grating parameters such as grating length and etching depth are investigated in this work as well. These parameters can be correlated to simulations and optimized for future processes. Emitters of identical construction can thus be realized in further device generations featuring optimized power-current-characteristics. It is also possible to integrate a grating section into the more complex device of a tapered laser. Such DBR-tapered lasers provide a spectrally stabilized and longitudinal single-mode emission with good beam quality so that world wide leading values for spectral radiance of 18,6 GW/sqcm/sr/nm can be calculated. Because the longitudinal mode filters are developed as a surface grating instead of a highly reflective but broad-band coating of the rear facet, a highly reflective and narrow-band DBR-section can be used. Accordingly, this thesis for the first time presents spectrally selective laser diodes which emit reliably high output power in continuous wave mode in the red spectral range. The accompanying chip-level integration offers potential series manufacturing as well as unprecedented miniaturization.en dc.identifier.uriurn:nbn:de:kobv:83-opus4-69776 dc.identifier.urihttps://depositonce.tu-berlin.de/handle/11303/4891 dc.identifier.urihttp://dx.doi.org/10.14279/depositonce-4594 dc.languageGermanen dc.language.isodeen dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/en dc.subject.ddc539 Moderne Physiken dc.subject.otherMonolithisches Oberflächengitterde dc.subject.otherRot-emittierende Diodenlaserde dc.subject.otherSchmale spektrale Linienbreitede dc.subject.otherSpektrale Selektivitätde dc.subject.otherDistributed bragg reflectoren dc.subject.otherMonolithic surface gratingen dc.subject.otherNarrow spectral linewidthen dc.subject.otherRed-emitting diode lasersen dc.subject.otherSpectral selectivityen dc.titleLongitudinale Modenfilter für Kantenemitter im roten Spektralbereichde dc.title.translatedLongitudinal mode filters for edge emitters in the red spectral rangeen dc.typeDoctoral Thesisen dc.type.versionpublishedVersionen tub.accessrights.dnbfree* tub.affiliationFak. 2 Mathematik und Naturwissenschaften::Inst. Festkörperphysikde tub.affiliation.facultyFak. 2 Mathematik und Naturwissenschaftende tub.affiliation.instituteInst. Festkörperphysikde tub.identifier.opus46977 tub.publisher.universityorinstitutionTechnische Universität Berlinen Files Original bundle Now showing 1 - 1 of 1 Loading… Thumbnail Image Name: feise_david.pdf Size: 19.43 MB Format: Adobe Portable Document Format Collections
Skip to main content Development of CNN models for the enteral feeding tube positioning assessment on a small scale data set Abstract Background Enteral nutrition through feeding tubes serves as the primary method of nutritional supplementation for patients unable to feed themselves. Plain radiographs are routinely used to confirm the position of the Nasoenteric feeding tubes the following insertion and before the commencement of tube feeds. Convolutional neural networks (CNNs) have shown encouraging results in assisting the tube positioning assessment. However, robust CNNs are often trained using large amounts of manually annotated data, which challenges applying CNNs on enteral feeding tube positioning assessment. Method We build a CNN model for feeding tube positioning assessment by pre-training the model under a weakly supervised fashion on large quantities of radiographs. Since most of the model was pre-trained, a small amount of labeled data is needed when fine-tuning the model for tube positioning assessment. We demonstrate the proposed method using a small dataset with 175 radiographs. Result The experimental result shows that the proposed model improves the area under the receiver operating characteristic curve (AUC) by up to 35.71% , from 0.56 to 0.76, and 14.49% on the accuracy, from 0.69 to 0.79 when compared with the no pre-trained method. The proposed method also has up to 40% less error when estimating its prediction confidence. Conclusion Our evaluation results show that the proposed model has a high prediction accuracy and a more accurate estimated prediction confidence when compared to the no pre-trained model and other baseline models. The proposed method can be potentially used for assessing the enteral tube positioning. It also provides a strong baseline for future studies. Peer Review reports Background Enteral nutrition through feeding tubes serves as the primary method of nutritional supplementation for patients unable to feed themselves. The position assessment of the Nasoenteric feeding tubes is essential following insertion and before the commencement of tube feeds to avoid potential complications [1]. A plain radiograph is typically performed to confirm the placement of the feeding tube [2]. The feeding tube position assessment is straightforward but costly and time-consuming. Timely interpretation of the radiographs remains a challenge affecting clinical decision to start tube feeds. Recently, artificial neural networks (ANNs) have shown great potential to be an effective tool to detect and diagnose medical problems [3,4,5]. As a data-driven approach in the concept of supervised learning, ANNs learn data features automatically from the given training set of samples and labels [6,7,8]. Deep convolutional neural networks (CNNs), as a subset of ANNs, have shown promising results in various medical imaging analysis tasks [9,10,11]. For instance, Esteva et al. applied CNN models to dermatoscopy images for skin cancer diagnosis and achieved performance on par with all participated human dermatologists. Ribli et al. used an rCNN-based [12] method for 2D mammograms classification that achieved 0.95 AUC for breast tumor classification [13]. Ying et al. proposed a cross-modality ANN model for Alzheimer’s disease (AD) diagnosis that used a CNN to evaluate head MRIs and a multilayer perceptron (MLP) model to analysis the single-nucleotide polymorphisms (SNPs) information from Genome-wide association study (GWAS) [14]. Their proposed model achieved a 0.935 AUC on AD diagnosis. However, robust CNNs commonly require large amounts of manually annotated data for training [15,16,17,18], such as ImageNet [19], which contains over one million images with labels. The prohibitively high annotation cost often presents a barrier to adopting modern CNN techniques in the medical imaging analysis tasks [20,21,22,23]. Transfer learning and pre-training are widely used in the medical imaging analysis that enables the training of CNN on small datasets [24,25,26]. In general, this includes three steps: first, a CNN model is pre-trained on a large dataset, such as ImageNet, for classification tasks; second, the feature extractor (i.e., the convolutional layers) of the pre-trained model is selected to be used as the backbone building block of another CNN model; third, the newly built CNN model is fine-tuned on a small medical imaging dataset for the specific purpose. Transfer learning from the ImageNet dataset to medical datasets has shown a promising result in improving the network performance on small medical datasets [24, 26,27,28], such a technique is also used to build enteral feeding tube positioning assessment models. For instance, Singh et al. transferred the ImageNet pre-trained model to the enteral feeding tube positioning assessment task and significantly improved the small training set [24]. However, an obvious domain gap exists between ImageNet images (i.e., natural images) and medical images, raising concerns about such a method [29]. We propose to use a novel pre-training method [30] to train CNN models on a small datasets for enteral feeding tube positioning assessment. Different from the early study [24], our method uses radiological imaging reports as weak supervision to pre-train the feature extractor on a large radiograph dataset before transfer learning is applied to build the feeding tube positioning assessment model. Radiological imaging reports are routinely collected in clinical practice and readily available in the medical record system. No additional manual labeling is required for pre-training of the proposed method. More importantly, the radiographs for pre-training are directly relevant to the enteral feeding tube positioning assessment task, mitigating the domain gap between natural imaging and radiographs posed by pre-training on the ImageNet dataset. Fig. 1 figure 1 An example of a radiograph (left) with the radiology report (right) from the MIMIC-CXR dataset [31] Method Model development We previously developed a general pre-training strategy, which used the radiology reports as weak supervision to pre-train a CNN model that improves the model performance on a given task [30]. This work extended the previous method to build an automatic enteral feeding tube positioning assessment network using a small training dataset. We assume two datasets, $X_P$ and $X_L$ ($\|X_P\| \gg \|X_L\|$), exist, where $X_P$ contains paired of radiographs and associated radiology reports and $X_L$ consists of labeled radiographs for enteral feeding tube positioning assessment. Our proposed network pre-trained the feature extractor of the enteral feeding tube positioning assessment model on $X_P$ directly without requiring manually annotated labels. The feature extractor, then, was fine-tuned on $X_L$ for the enteral feeding tube positioning assessment. Figure 1 shows an example of a radiograph and the corresponding radiology report. Pre-training feature extractor via radiograph-report matching We pre-trained the feature extractor of the enteral feeding tube positioning assessment model through a radiograph-report matching network (Fig. 2), containing a textual report processing branch (Fig. 2a), a radiograph processing branch (Fig. 2b), and a contrastive learning module (Fig. 2c). The two branches worked simultaneously in parallel. The network took a radiology report and radiograph pair as input and predicted whether they were a natural match. Since label (i.e., match or don’t match) is known, no manual annotation will be required. This weakly supervised pre-training approach transfers the rich information in reports to the radiograph feature extractor without requiring manually labeled data. Fig. 2 figure 2 Illustration of the weakly supervised pre-training approach Specifically, the textual report processing branch (Fig. 2a) took a radiology report as input and (1) passed the report through a pre-trained BERT (Bidirectional Encoder Representations from Transformers) [32] encoder and a $1\times 1$ convolutional (Conv) layer to convert the natural language in the report to numerical embeddings, i.e., a sequence of numbers that can be processed by computer algorithms, (2) reduced the dimensionality of the embeddings by applying a global average pooling (GAP) operation, and (3) projected the embeddings to a latent feature space by a fully connected (FC) layer. The output of the textual report processing branch was a feature vector that represented the report in latent space. Meanwhile, the radiograph processing branch (Fig. 2b) took a radiograph as input and passed it through a ResNet-18 [15] feature extractor. The generated feature map was then passed through a Conv layer with $1\times 1$ kernels to transfer the pre-trained features to task-specific features. After that, an FC layer was used to embed the radiograph feature map to the latent space, which is the same as the textual report features. The output of the radiograph processing branch was a feature vector in the latent space that represented the input radiograph. Next, the radiograph-report matching network was trained in a contrastive manner via the contrastive learning module (Fig. 2c). A shallow CNN classifier was added on top of the two branches that takes the absolute difference between the two feature vectors as input and ouputs whether the two feature vectors belonged to the same example. Mathematically, the radiograph-report matching network could be written as: $$\begin{aligned} h_{\theta _p}(x^{i}) = h_{\theta _{cls}}(\|h_{\theta _t}(x_t^{i}) - h_{\theta _r}(x_r^{i})\|). \end{aligned}$$ (1) where $x^{i}=\{x_t^{i}, x_r^{i}\}$ was a pair of a textual radiology report, $x_t^{i}$, and a radiograph, $x_r^{i}$ from $X_P$. Note that $x_t^{i}$ and $x_r^{i}$ may or may not match. The network $h_{\theta _p}(\cdot )$ predicted the probability of the input pair being a natural match. The $h_{\theta _{cls}}(\cdot )$ was the contrastive learning module, $h_{\theta _t}(\cdot )$ was the textual report processing branch, and $h_{\theta _r}(\cdot )$ was the radiograph processing branch. Binary cross-entropy loss was used to train the text-image matching network. The input of the radiograph-report matching network was a radiology report and radiograph pair. A label was naturally assigned to each radiograph-report shwoing whether are from the same imaging event. A true pair meant the report describes the radiograph naturally; otherwise, it was a false pair. CNN for enteral feeding tube Positioning assessment Fig. 3 figure 3 The enteral feeding tube positioning assessment network The enteral feeding tube positioning assessment model was trained by fine-tuning the feature extractor in the radiograph processing branch, $h_{\theta _r}(\cdot )$, of the pre-trained network. The process was straightforward and illustrated in Fig. 3. The Conv layers in $h_{\theta _r}(\cdot )$ were used as the feature extractor in the enteral feeding tube positioning assessment model. A Conv layer and two FC layers were added on top of the feature extractor to build the classification model for the enteral feeding tube positioning assessment network, $h_{\theta }(\cdot )$. The $h_{\theta }(\cdot )$ took radiographs from $X_L$ and predicted the probability of the enteral feeding tube positions being satisfied. Since the feature extractor was pre-trained using a larger dataset set from the same domain, we only need to optimize the $h_{\theta }(\cdot )$ from scratch that may use significantly reduce the need for the total number of training instances. Enteral feeding tube positioning dataset A dataset containing plain radiographs of 175 patients was retrospectively retrieved at a comprehensive tertiary academic medical center. All the images were inspected by a board-certified abdominal radiologist with more than 10 years of experience and a trainee. The dataset included 63 images where the enteral feeding tube positioning was unsatisfactory, and 112 images with a satisfying position. This retrospective study was approved by the Institutional Review Boards of the University of Kentucky. The pixel values of radiographs were converted to the range of 0-255 using a window of 0-2750. The images were resized to $256\times 256$ and equally split into five folds for a fivefold cross-testing. Real-time data augmentation for the combination of a random horizontal flip and rotation between 0 and 20 degrees was applied to the training data. Model evaluation Compared models We compared the proposed model with the CNN models trained using four different pre-training strategies: (a) a CNN model without pre-training (denoted as No Pre-Train), (b) a CNN model pre-trained on ImageNet [19] (denoted as ImageNet), (c) a CNN model pre-trained using Compare to Learning [29], a state-of-the-art self-supervised pre-training model for 2D medical images (denoted as C2L), and (d) a true random CNN model (denoted as Random). All models have the same architecture. The No Pre-Train model was a typical CNN model trained using the enteral feeding tube dataset only. No pre-training strategy was applied. All the weights of this model were randomly initialized before the training. The ImageNet model was a CNN model that pre-trained on the ImageNet dataset, a natural imaging dataset containing over one million images of 1000 classes. Such a pre-training method is well-accepted and widely used in the medical imaging domain [26,27,28], which was also used in [24], an early study of enteral feeding tube positioning assessment using CNN models. The model was trained on the ImageNet dataset for a classification task and was fine-tuned using the enteral feeding tube dataset under the same approach of Sect. 2.1.2. The C2L model was pre-trained using Comparing to Learn [29] on the MIMIC-CXR dataset [31] that was a self-supervised, pre-training method that was proposed for medical imaging analysis. The method pre-trained a feature extractor on MIMIC-CXR, containing 227, 835 radiographic studies of 64, 588 patients that including 368, 948 chest radiographs and the associated radiology reports. The model, then, was fine-tuned using the enteral feeding tube dataset under the same approach of Sect. 2.1.2. The Random model was a CNN model with randomly initialized weights. The model was not trained with any data samples. The model performs random guessing for any input examples. The proposed model was pre-trained using [30] that was proposed by our previous study. Specifically, the feature extractor of the proposed method was pre-trained on MIMIC-CXR for radiograph-report matching tasks. The detailed pre-training setup of the proposed method was described in [30]. After the feature extractor was pre-trained, the network was fine-tuned using the enteral feeding tube dataset under the same approach of Sect. 2.1.2. No radiology reports were needed for fine-tuning or testing the enteral feeding tube positioning assessment model. All the compared models were trained for five trials with a fivefold cross-testing strategy. We used three folds for training, one for validation, and one for testing. We repeated this process until all folds were tested. The validation fold is used to select the best checkpoint of the model. Then, the selected checkpoint is used to test the model on the testing fold. The Cyclic learning rate [33] between $10^{-4}$ and $10^{-2}$, Adam optimizer [34], and binary cross-entropy loss were used for the enteral feeding tube dataset training or fine-tuning. All the models were trained for 100 epochs. We used Python as the programming language and PyTorch [35] as the scientific computing library to conduct the evaluation. For the ImageNet pre-trained model, we loaded the PyTorch pre-trained weights directly in to the model. The training was performed on a GPU cluster that has a combination of 120 Nividia P100 and V100 GPU cards. However, only one GPU card was used for the training at the same time. Evaluation metrics Four evaluation metrics were used in this study, namely the AUC, F1 score, accuracy, and the expected calibration error (ECE) [36]. The AUC, F1 score, and accuracy were used to evaluate models’ performance in making accurate predictions. All three metrics were bound between 0 to 1. A higher number indicated better performance. The ECE was used to measure neural network calibration error, i.e., how accurately the network estimates its prediction confidence, with a smaller value indicating a more accurate representation of its prediction confidence. A perfectly calibrated neural network has a 0 ECE. We defined the accuracy, AUC, and F1 score following common practice. The ECE was defined as the same as [36, 37] by partitioning predictions into M bins and taking a weighted average of the difference of accuracy and confidence for each bin. More specifically, we first grouped all the samples into M interval bins according to the predicted probability. Then, let $B_m$ be the set of indices of samples whose predicted confidence falls into the interval $I_m=(\frac{m-1}{M}, \frac{m}{M}]$, $m \in M$. The ECE can be calculated as: $$\begin{aligned} \text {ECE} = \sum _{m=1}^{M} \frac{\|B_m\|}{n} \left\| \frac{1}{\|B_m\|} \sum _{i\in B_m} 1\cdot ({\hat{y}}^i = y^i) - \frac{1}{\|B_m\|} \sum _{i\in B_m} {\hat{p}}^i\right\| , \end{aligned}$$ (2) where n was the number of samples, ${\hat{y}}^i$ and $y^i$ were the predicted and ground-truth label for sample i, ${\hat{p}}^i$ was the confidence of sample i, $\frac{1}{\|B_m\|} \sum _{i\in B_m} 1\cdot ({\hat{y}}^i = y^i)$ was the accuracy of $B_m$, and $\frac{1}{\|B_m\|} \sum _{i\in B_m} {\hat{p}}^i$ calculated the average predicted confidence of $B_m$. Model interpretation Integrated Gradients attribution mask (IG) [38] and occlusion sensitivity testing map (OCC) [39] are used as visualization methods to understand how predictions are made by the proposed model. IG is an interpretability technique for CNN models that visualize the important features that contribute to the model’s prediction. Higher values in an IG attribution mask indicate more important features in the decision-making process. OCC is a technique for understanding which parts of an image are most important for a CNN classification. The higher values in an OCC map indicate more important areas for the image during the CNN classification procedure. Results Table 1 presents the detailed evaluation result of the five models with the mean score and the 95% confidence interval of each evaluation metrics over the five trials. From the table, we can see that the proposed method has the best overall performance, which has better or comparable performance to other compared models in all settings. The C2L model achieves the second-best overall performance on prediction accuracy evaluation metrics but performs poorly on network calibration. The ImageNet model wins third place. The No Pre-Train model has a better performance than the Random model and gets the fourth place. The Random model performs the worst. Table 1 Performance of each model (mean, $95\%$ confidence interval) The table shows that the Random model has a 0.49 AUC, which is essentially random guessing. The No Pre-Train model yields a 0.56 AUC. The ImageNet improves the number to 0.67. The C2L further improves it to 0.73. The proposed method has the highest AUC score of 0.76. For the F1 score, the Random model, No Pre-Train model, and ImageNet model have F1 scores that are either close to 0.5 or lower, with the worse of 0.32 for the No Pre-Train model. The proposed method can improve the F1 score by 100% when compared with the No Pre-Train model, from 0.32 to 0.64, which is also 20.75% and 3.13% higher than the ImageNet and C2L models, respectively. The C2L model has the highest accuracy, 79%, which is approximately 1% higher than the proposed model (78%). The No Pre-Train model and ImageNet model have 74% and 69% accuracy, respectively. The Random model has 49% accuracy. The C2L model has the highest calibration error, 0.15 ECE, which is 66.67% higher than the proposed method (0.09). Both the No Pre-Train model and ImageNet model have 0.12 ECE. The Random model has 0.11 ECE. Fig. 4 figure 4 Visualization of two correct predictions Figures 4 and 5 show four examples of the enteral feeding tube positioning assessment results and corresponding Integrated Gradients attribution mask (IG) and occlusion sensitivity testing map (OCC). Figure 4 shows that for the correctly predicted cases, the IG and the OCC highlight the areas that critical to assessing the enteral tube positioning, while Fig. 5 shows that for the failure cases, the network was focusing on the areas that were less important to assessing the enteral tube positioning. Fig. 5 figure 5 Visualization of two wrong predictions Discussion The hypothesis of our proposed method transfers natural language to image features of the architecture during the radiograph-report matching stage. The radiograph-report matching process pretrains the radiograph feature learning component parameters to the extent that it needs fewer supervised instances for the feeding tube positioning assessment task. The result reveals that the Random model has an essential random performance (0.49 AUC and 48% accuracy) that sets the bottom-line performance of the enteral feeding tube positioning assessment task on this given dataset. The No Pre-Train improves the AUC and accuracy to 0.56 and 69%, respectively, which indicates the model did learn something about assessing the tube positioning from the training set. However, the low F1 score of the No Pre-Train (0.32) may suggest that the model is extremely biased to one class when making the prediction. This unexpected behavior is likely due to the small size of the training data. It is widely accepted that transfer learning helps to improve model performance on a small dataset. As expected, all three transfer learning models have a better performance than the No Pre-Train model. The ImageNet model improves the AUC to 0.67, the C2L model pushes the number to 0.73, and the proposed method achieved a 0.76 AUC. Though both the F1 score and accuracy are also improved by the three transfer learning models, the larger gap between the F1 score and accuracy of the ImageNet and C2L models may indicate those two models also favor one class when making the decision, especially for the ImageNet model. Ideally, a predicting model should be able to reflect the uncertainty or the confidence of its prediction accurately. Otherwise, it may be problematic. For instance, given k predictions with average prediction confidence of c ($c \le 1.0$), we could expect $\approx k\times c$ correct predictions or an automatic $\approx c$% accuracy. However, the average prediction confidence often does not match the accuracy for the modern deep neural networks [40,41,42]. ECE is the common metric to evaluate neural network calibration error. We believe accurately estimated CNN prediction confidence is extremely important to automatic medical imaging analysis tools because an automated method that achieves high accuracy but captures prediction confidence inaccurately could lead to significant treatment errors [43]. Table 1 shows that the proposed model has the lowest ECE (0.09), which is 40% less than the C2L model and 25% less than the ImageNet model. One reasonable explanation is the precise guidance of the proposed method helps reduce the ECE on the downstream application. The proposed method is pre-trained on radiographs with radiology reports that are more relevant to enteral feeding tube positioning assessment than ImageNet pre-training, which uses natural images to pre-train the network weights. Though the C2L is also pre-trained on radiographs, the method uses a self-supervised strategy and provides an open ending to some degree. By observing the correctly and wrongly predicted cases, we notice that most of the wrongly predicted cases are more challenging than the correctly predicted cases, even for human experts. For example, the first case in Fig. 5 contains other wires, which confuse the model. The IG and OCC show that the model pays more attention to those external wires when making the decision. The second case in Fig. 5 contains stronger noisy patterns, which make the images harder to read. A direct approach to improve the performance of challenging cases is to obtain more training data of challenging cases. However, this may not be easy due to the high cost of data collection. Thus, we plan to tackle this scenario from the algorithmic perspective by adding weights to challenging cases and using boosting strategies. One limitation of this study is the lack of a standalone dataset for testing since the dataset is small. We apply a fivefold cross-testing strategy to generate a more objective testing result. Such a strategy is widely used in other specific imaging domains [44, 45]. It may be more objective than regular fivefold cross-validation because the results are based on the unseen testing set, not the validation set. A multi-site, large-scale evaluation may still be needed for further testing before using the proposed method in clinical practice. Conclusion We propose a novel enteral feeding tube positioning assessment network, which can be trained using a small-scale dataset. 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Author information Authors and Affiliations Authors Contributions Jie Zhang supervised the whole project. Gongbo Liang and Liangliang Liu conducted the experiments and neural network training. Gongbo Liang and Jie Zhang wrote the manuscript. Halemane Ganesh supervised the project as an experienced radiologist. Halemane Ganesh and Dylan Steffey provided the data and verified the results. Nathan Jacob contributed to technical method development. All authors read the manuscript and contributed to the manuscript revision. All authors read and approved the final manuscript. Corresponding authors Correspondence to Gongbo Liang or Jie Zhang. Ethics declarations Ethics approval and consent to participate This retrospective study was approved by the University of Kentucky institutional review board, with a waiver of informed consent and compliant with the Health Insurance Portability and Accountability Act. All methods were carried out in accordance with relevant guidelines and regulations. Consent for publication Not applicable. Competing interests The authors declare that they have no competing interests. Additional information Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Rights and permissions Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Reprints and permissions About this article Check for updates. Verify currency and authenticity via CrossMark Cite this article Liang, G., Ganesh, H., Steffe, D. et al. Development of CNN models for the enteral feeding tube positioning assessment on a small scale data set. BMC Med Imaging 22, 52 (2022). https://doi.org/10.1186/s12880-022-00766-w Download citation • Received: • Accepted: • Published: • DOI: https://doi.org/10.1186/s12880-022-00766-w Keywords
Send email with Free (French host) SMTP I reach to make it works some weeks ago, but it doesn’t anymore (I don’t remember to have change my settings). Here they are : WM_SocketSMTP.Address = “smtp.free.fr WM_SocketSMTP.Port = 587 WM_SocketSMTP.SSLConnectionType = SSLSocket.SSLConnectionTypes.SSLv23 WM_SocketSMTP.SMTPConnectionMode = 1 WM_SocketSMTP.SSLEnabled = True WM_SocketSMTP.Username = “tom.25” ’ my email is @free.fr WM_SocketSMTP.Password = “MyPassword” In this case, the email never go, the program wait and wait until I cancel. If I do : WM_SocketSMTP.SSLEnabled = False Then the email seems to have been sent but no. As if I do : WM_SocketSMTP.Port = 465 It seems to have been sent but neither, no. :confused: My settings are the same as in Apple Mail (where I can send email) except authentification which is (if I translate in English) “Answer to chalenge MD5” Note I hesitate to write inFrench and post in the French sub-forum. But maybe I do something wrong. I have an Address Orange and it works, except the port is 465. as tu activé l’envoi de mail sécurisé sur le portail de ton compte free ? as tu essayé d’envoyer un mail sans aucun mot de passe ni identifiant (si tu envoie depuis une freebox il n’y a pas besoin d’identifiant ni de mot de passe) You should get at least an error or a timeout. The part where you make your email is missing from the code. Try with the CURL plugin from MBS or the Chilkat plugin. The plugins have much better error reporting. Merci jean-Yves, oui c’est bien activé. j’ai sur ma page Free : Votre login "tom.25" est déjà autorisé à faire du SMTP authentifié sur "smtp.free.fr". Seules les méthodes d'authentification avec mot de passe chiffré sont acceptées. Par exemple l'utilisation du SSL (sur le port 465) ou du 'MD5 Challenge-Response' (sur le port 587). Pour désactiver cette fonctionnalité, cliquez ici: Si j’essaye sans identifiant, mon programme (Xojo) me retourne immédiatement l’erreur : Server Error 554554 5.7.1 <unknown[109.221.31.99]:53928>: Client host rejected: Access denied Je suis chez Sosh (Orange), j’ai une LiveBox et non une Freebox. HS : Et avant hier ils ont posé la fibre devant chez moi. Ca arrive :slight_smile: Thanks Beatrix, no Xojo doesn’t display any error. This morning I received one of the emails I sent me yesterday during my test, but which settings it was ??? :confused: I prefer to stay with Xojo code, I know MBS pluggins are wonderfull but I can’t buy them. And learn to use another pluggin is too long for me. I will send my email with another host (orange) if I can’t with Free host. Beatrix, I didn’t paste all my code but it works when I use other host. I just see that the instruction : WM_SocketSMTP.SMTPConnectionMode = 1 is API1 and is not shown as deprecated. It should be : WM_SocketSMTP.SMTPConnectionType = SMTPSecureSocket.SMTPConnectionTypes.SSLTLS I will fill a FeedBack. (it does not resolve my problem). I didn’t paste all my code but it works when I use other host. It works with another host ? Usually, you can only use the SMTP server from your current provider, i.e. you can use smtp.free only if you are currently connected to the internet via a Free account. If you, let’s say your phone, is connected to another network like Orange or SFR, then the server smtp.free will reject your connection. not if you authenticate
What Mercedes is equivalent to the SQ5? 1. Home page 2. F.A.Q What Mercedes is equivalent to the SQ5? If you’ve been asking yourself “what Mercedes is equivalent to the SQ5?” then you’ve come to the right place. The Mercedes-Benz GLC43 AMG is the luxury vehicle closest to the Audi SQ5 and has many similar features. The GLC43 AMG offers a 3.0-Liter V-6 Biturbo engine that produces 385-hp and 384 lb-ft of torque, giving it an impressive top speed of 156 mph. This Mercedes also offers a nine-speed automatic transmission and a 4MATIC all-wheel drive system, allowing for balanced performance in a variety of driving circumstances. The exterior of the GLC43 AMG is also quite pleasing, with its sleek and stylish design. It comes with 19-inch black wheels, a panoramic sunroof, and a set of LED headlights and taillights. Inside, it has a comfortable and luxurious interior that is filled with leather upholstery, wood trim, and a 12.3-inch infotainment system. There is also a plethora of safety features, such as adaptive cruise control, lane keep assist, and a 360-degree camera system. The GLC43 AMG is a great option for those in the market for a luxury crossover that is comparable to the SQ5. It is priced at around $56,000, making it slightly more expensive than the SQ5, but it also offers more features and a higher level of performance. Overall, the GLC43 AMG is a great option for those looking for a luxury vehicle similar to the Audi SQ5. It offers a powerful engine, a stylish exterior, and a luxurious interior, as well as a plethora of safety features and advanced technologies. If you’re looking for a luxury vehicle that is similar to the SQ5, then the GLC43 AMG is a great choice. What Mercedes is equivalent to the SQ5? Comparing the Mercedes GLC Coupe to the Audi SQ5 If you’re looking for a luxurious SUV and you’re torn between the Mercedes GLC Coupe and the Audi SQ5, it can be difficult to make a decision. Both cars have plenty of power, high-end features, and superior comfort. In this article, we’ll compare the two cars to help you make an informed decision. The Mercedes GLC Coupe is a luxury SUV with a sleek exterior design and a powerful engine. It has a 3.0-liter V6 engine that produces 362 horsepower and 369 lb-ft of torque. The engine is mated to a 9-speed automatic transmission, and the GLC Coupe can reach a top speed of 155 mph. It also offers standard features like a 10.25-inch display, active brake assist, and an air suspension. The Audi SQ5 is the performance variant of the Audi Q5. It has a 3.0-liter turbocharged V6 engine that produces 349 horsepower and 369 lb-ft of torque. It’s mated to an 8-speed automatic transmission and can reach a top speed of 155 mph. The SQ5 also comes with standard features like a 12.3-inch display, active lane assist, and adaptive suspension. When it comes to price, the Mercedes GLC Coupe and the Audi SQ5 are pretty comparable. The GLC Coupe starts at around $50,000, while the SQ5 starts at around $54,000. Both cars also offer a variety of packages and upgrades that can increase the price significantly. In terms of performance, the Audi SQ5 has the edge. It has more power and can reach higher speeds. If you’re looking for a luxury SUV that offers superior performance, the SQ5 is the better option. Both cars offer plenty of modern features, but the Audi SQ5 has a few more options. It has a higher-resolution display, active lane assist, and adaptive suspension. The GLC Coupe does have some features that are not available on the SQ5, like active brake assist and an air suspension. To summarize, the Mercedes GLC Coupe and the Audi SQ5 are both excellent luxury SUVs. The GLC Coupe is a great choice if you’re looking for a sleek and comfortable ride, while the SQ5 offers more power and performance. Both cars offer plenty of features and modern amenities, but the SQ5 has a few more options. AttributeMercedes GLC CoupeAudi SQ5 Engine3.0-liter V63.0-liter turbocharged V6 Transmission9-speed automatic8-speed automatic Top Speed155 mph155 mph Display10.25-inch12.3-inch Price$50,000$54,000 What Mercedes is equivalent to the SQ5? 2 What Features Make the Mercedes GLC Coupe a Versatile Option for Driving Comfort and Luxury? The Mercedes GLC Coupe is one of the most versatile luxury cars on the market. It is a perfect combination of driving comfort and luxurious features. It is the perfect option for those who want to enjoy the best of both worlds. This car features a powerful 3.0-litre V6 engine that provides plenty of power and torque for any situation. The seven-speed automatic transmission is smooth and efficient, making it a great option for highway cruising. It also features a range of driver-assistance technologies, such as Active Brake Assist, Blind Spot Assist, and Crosswind Assist. As a result, it is incredibly safe and secure on the roads. The Mercedes GLC Coupe also offers a wide range of comfort and convenience features. It comes standard with a 12.3-inch central display, a COMAND infotainment system with touchpad controller, heated seats, and more. As a result, it is a great option for long-distance trips or daily commutes. It also has plenty of room in the cabin for passengers and cargo, so you can bring along all your essentials. The Mercedes GLC Coupe is also incredibly luxurious, with a plethora of features that make it a great option for those who love to drive in style. It has a range of premium materials, such as Nappa leather and real wood accents. It also comes with a range of advanced safety features, such as adaptive cruise control, active lane keeping assist, and more. When it comes to luxury, the Mercedes GLC Coupe is a great option. It is available in a range of trims and packages, so you can find the perfect car to fit your needs. Prices start at around $48,000, which is a great deal for a car with so much to offer. For those looking for a versatile luxury car, the Mercedes GLC Coupe is a great option. It has a powerful engine, plenty of advanced features, and plenty of room for passengers and cargo. It is also incredibly luxurious, and it has a price tag that won’t break the bank. All in all, it is the perfect combination of driving comfort and luxury. What Mercedes is equivalent to the SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What is the model of Mercedes that is most like the SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What SUV from Mercedes is closest to the Audi SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. Can you compare the SQ5 to a Mercedes SUV? Yes, the Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What Mercedes is most similar to the Audi SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What is the Mercedes equivalent to the Audi SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What model of Mercedes has features most like the SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. Which Mercedes model is the equivalent of the SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What is the Mercedes model that has the same features as the SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. What is the closest Mercedes model to the Audi SQ5? The Mercedes-AMG GLC 43 4MATIC is the model that is most comparable to the Audi SQ5. Your email address will not be published. Required fields are marked *
Advertisement Upgrade to remove ads The Special Senses of the Human Body Special senses -have receptors strategically placed in unique organs -include: olfaction, taste, visual system, hearing & balance Olfaction key points -7 primary odors now recognized, but average person can recognize 4000 different odors. perceived by olfactory epithelium. -dendrites of olfactory neurons have enlarged ends (olfactory vesicles) -cilia (olfactory hairs) of olfactory neuron embedded in mucus. odorants dissolve in mucus. Olfaction key points 2 -odorants attach to receptors, cilia depolarize & initiate action potentials in olfactory neurons. one receptor may respond to more than one type of odor. -olfactory epithelium replaced as it wears down. olfactory neurons replaced by basal cells every 2 months. unique: most neurons are permanent cells (aren't replaced if they die). Neuronal Pathways of Olfaction -olfactory sensory pathway: olfactory neurons (bipolar) in the olfactory epithelium pass through cribiform plate to olfactory bulbs & synapse w/ tufted cells or mitral cells -these extend to olfactory tract & synapse w/ association neurons -association neurons also receive input from brain, so info can be modified before it reaches the brain. Neuronal Pathways of Olfaction 2 -info goes to olfactory cortex of frontal lobe w/out going through thalamus (only major sense that does not go through thalamus). -3 regions in frontal lobe affect conscious perception of smell & interact w/ limbic system -lateral, medial, & intermediate olfactory areas Lateral olfactory area conscious perception of smell Medial olfactory area visceral & emotional reactions to odors Intermediate olfactory area effect modification of incoming info Taste Gustatory Taste buds -the sensory structures that detect taste -includes supporting cells surrounding taste (gustatory) cells -taste cells have microvilli (gustatory hairs) extending into taste pores -replaced about every 10 days Papillae -specialized regions on the tongue -types: filiform, vallate, fungiform, & foliate Filiform filament-shaped; provide a rough surface for food manipulation Vallate largest, least numerous. 8-12 in V along border between anterior & posterior parts of tongue. have taste buds. Fungiform mushroom-shaped. scattered irregularly over the superior surface of tongue. look like small red dots interspersed among the filiform. have taste buds. Foliate leaf-shaped. in folds on the sides of the tongue. contain most sensitive taste buds. decrease in number w/ age. Tastants -substances which are dissolved in saliva, enter the taste pores. -by various mechanism (depending on the taste), tastants cause the taste cells to depolarize. Taste types sour, salty, bitter, sweet, umami Sour most sensitive receptors on lateral aspects of the tongue. H+ ion of acids cause depolarization. Salty most sensitive receptors on tip of tongue. shares lowest sensitivity w/ sweet. anything w/ Na+ causes depolarization. Bitter most sensitive receptors on posterior aspect. highest sensitivity. sensation produced by alkaloids, which are toxic. Sweet most sensitive receptors on tip of tongue. shares lowest sensitivity w/ salty. sugars, some carbohydrates, & some proteins (NutraSweet: aspartame). Umami "Savory"; scattered sensitivity. caused by amino acids such as glutamate, binding to receptors Taste 2 -texture affects the perception of taste -temperature affects taste perception -very rapid adaptation, both at level of taste bud & within CNS -taste influenced by olfaction -different tastes have different thresholds w/ bitter being the taste to which we are most sensitive. many alkaloids (bitter) are poisonous The Vertebrate Eye cornea, choroid, iris, aqueous humor, vitreous humor, lens, retina, macula lutea, fovea, conjunctiva cornea transparent, admits light choroid is the vascular layer of the eye, provides oxygen & nourishment to the outer layers of the retina iris -behind the cornea -controls diameter of pupil -regulates amount of light that strikes the lens aqueous humor a clear, gelatinous fluid in front chamber of eye. maintains the intraocular pressure & inflates the globe of the eye. vitreous humor clear gel that fills the space between the lens & the retina of the eyeball; helps hold the eye in place lens focuses image on the retina retina -lines the back of the eye -photoreceptors and neurons integrate info detected by photoreceptors macula lutea is an oval-shaped highly pigmented yellow spot near the center of the retina fovea found in the macula lutea; provides highest resolution vision; contains a lot of cone cells (for color vision) conjunctiva thin transparent mucous membrane three tunics (tunic=layer): fibrous: sclera & cornea (outer layer) vascular: choroid, ciliary body, iris (middle) nervous: retina fibrous tunic contains sclera & cornea sclera white outer layer. maintains shape, protects internal structures, provides muscle attachment point, continuous w/ cornea cornea connective tissue matrix containing collagen, elastic fibers, & proteoglycans -avascular, transparent, allows light to enter eye; bends & refracts light vascular tunic middle layer. contains most of blood vessels of eye: branches off the internal carotid arteries. contains melanin. -contains iris, ciliary body, & choroid iris colored part of eye. controls light entering pupil. smooth muscle determines size of pupil. -sphincter pupillae: circular smooth muscle; contraction=decrease pupil size (or constricts) -dilator pupillae: radial smooth muscle; contraction=dilation of pupil ciliary body produces aqueous humor that fills anterior chamber -ciliary muscles: smooth muscle that controls lens shape choroid associated w/ sclera. very thin, pigmented. is vascularized. nervous tunic 2 layers: pigmented & sensory retina pigmented retina outer, pigmented layer. pigment of this layer & choroid help to separate sensory cells & reduce light scattering. sensory retina inner layer of rod & cone cells sensitive to light anterior compartment -anterior to lens; filled w/ aqueous humor -contains 3 chambers of eye 3 chambers of eye anterior, posterior & vitreous anterior chamber between cornea & iris posterior chamber -between iris & lens -helps maintain intraocular pressure; supplies nutrients to structures bathed by it; contributes to refraction of light glaucoma abnormal increase in intraocular pressure vitreous chamber posterior to lens. filled w/ jelly-like vitreous humor. helps maintain intraocular pressure, holds lens & retina in place, refracts light. the lens -held by suspensory ligaments attached to ciliary muscles. changes shape as ciliary muscles contract & relax -transparent, biconvex visible light portion of electromagnetic spectrum detected by human eye refraction bending of light convergence light striking a convex surface focal point -point where light rays converge & cross -lens changes shape causing adjustment of focal point on the retina focusing causing light to converge near point of vision closer than 20 feet. changes occur in lens, size of pupil, & distance between pupils. accommodation ciliary muscles contract due to parasympathetic input via cranial nerve III. pulls choroid toward lens reducing tension on suspensory ligaments. lens becomes more spherical, greater refraction of light. pupil constriction varies depth of focus convergence as objects move close to the eye, eyes are rotated medially. reflex contraction of the medial rectus muscles. how the eye processes light involves several cells, especially photoreceptor cells called rods & cones Amacrine & horizontal cells help integrate & regulate the input from multiple photoreceptor cells (eg. horizontal cells are responsible for allowing eyes to adjust to see well under both bright & dim light conditions) Rods specialized for detection of low-intensity light Cones specialized for detecting light of different wavelengths (colors) The Photoreceptor cells of the Eye (called rods & cones)... -contain photoreceptive molecules that -absorb energy of light -generate changes in membrane potential -in rods, photopigment is rhodopsin -in cones, photopigment is iodopsin Photoreceptive Molecules (such as rhodopsin) -absorb light in photoreceptor cells -consist of retinal combined w/ an opsin protein -there are 3 types of photopigments in cones (called photopsins) -photopigments are found in the discs of photoreceptor cells (that is, the rods & cones) Rhodopsin= retinal+opsin=photoreceptive molecule found in rods; is a G-protein coupled receptor In the rod photoreceptor cell: steps 1-4 1. cis-Retinal absorbs light; is converted to trans-retinal 2. the protein segment (opsin) of rhodopsin is activated, triggering activation of G protein transducin 3. the activated G protein activates phosphodiesterase 4. activated phosphodiesterase breaks down cGMP to 5'-GMP, which then detaches from the Na+ channel In rod photoreceptor cell: steps 5-7 5. loss of cGMP closes Na channel 6. membrane hyperpolarizes & reduces neurotransmitter release 7. when glutamate is not released (or reduced) from the rod cell, the bipolar cell is no longer inhibited & can now stimulate ganglion cells, produce APs, send signal to brain Processing visual info -in dark, rods & cones are depolarized & continually release glutamate -some bipolar cells depolarize in response to glutamate, others hyperpolarize (depending on type of receptor present on bipolar cell) -when light hits rods & cones, they hyperpolarize (b/c Na+ channel that was originally open in dark, that allowed Na+ to enter & depolarize cell, has been closed) -this shuts off release of glutamate -some bipolar cells will now be no longer inhibited by glutamate. they release neurotransmitters which stimulate ganglionic cells to generate APs to the brain G-protein-linked receptors -rhodopsin is a G-protein linked receptor found on membrane of rod photoreceptor cells -utilizes cyclic GMP: -concentration of GMP controlled by: -synthesis by guanylyl cyclase, -degradation by cyclic GMP phosphodiesterase -light activates rhodopsin, causing phosphodiesterase to convert to cGMP to 5' GMP -this closes Na+ channel on rod cell -glutamate is no longer released from rod cell -bipolar cell no longer inhibited & can send APs to ganglionic cells to brain Rods light & dark adaptation -in bright light, more rhodopsin broken down to Vitamin A, protecting eye & making it less sensitive to light -in darker conditions, more rhodopsin produced so eye is more sensitive to light -takes eyes a while to accommodate when going from dark to light & vice versa b/c of these chemical changes that must occur Pupils light & dark adaptation constriction in bright light; dilation in dim light Cones responsible for color vision & visual acuity -numerous in fovea & macula lutea; fewer over rest of retina -visual pigment is iodopsin: 3 types that respond to blue, red & green light -overlap in response to light, thus interpretations of gradation of color possible: several millions Receptive Fields -area from which a ganglion cell receives input -roughly circular w/ receptive field center -2 types of receptive fields: on-center & off-center cells -interneurons present in inner layers & modify signal b/f signal leaves retina. enhance borders & contours, increasing intensity at borders on-center ganglion cells generate more action potentials when light is directed onto the receptive field. respond to intensity of light off-center cells more action potentials when light is off or when light does not hit center of field. respond to contrasts in light. How is location of a visual stimulus encoded in nervous system? I. Comparing input: -from 2 eyes gives: left/right location, & distance (or depth perception). -when eyes move up & down gives: up/down location II. Map-like projection from retina to cortex relays the left/right, up/down location, & distance (depth perception) info Visual Fields -close 1 eye. everything you can see w/ your open eye=the visual field for that eye -visual fields of each eye partially overlap -region of overlap=area of binocular vision -see same object w/ both eyes. thus image of object reaches retina of one eye at a slightly different angle from that of the other- results in depth perception. vision info in a visual field (eg, "right" visual field or "left" visual field) is processed in the opposite side of the brain Processed signal... is sent via optic nerve through lateral geniculate nuclei to visual cortex. Note: visual field to left is viewed by right side of both eyes & is processed in right visual cortex (& vice versa) Myopia nearsightedness -focal point too near lens, image focused in front of retina Hyperopia farsightedness -image focused behind retina Presbyopia degeneration of accommodation, corrected by reading glasses Astigmatism cornea or lens not uniformly curved Strabismus lack of parallelism of light paths through eyes Retinal detachment can result in complete blindness Glaucoma increased intraocular pressure by aqueous humor buildup Cataract clouding of lens Macular degeneration common in older people, loss in acute vision Diabetes dysfunction of peripheral circulation 3 divisions of ear external, middle, & inner ear external ear hearing. terminates at eardrum (tympanic membrane). includes auricle & external auditory canal middle ear hearing. air-filled space containing auditory ossicles. inner ear hearing & balance. interconnecting fluid-filled tunnels & chambers within the temporal bone external ear 2 -auricle or pinna: elastic cartilage covered w/ skin -external auditory canal: lined w/ hairs & ceruminous glands. produce cerumen. -tympanic membrane: -thin membrane of 2 layers of epithelium w/ connective tissue btwn -sound waves cause it to vibrate -border btwn external & middle ear middle ear 2 -separated from inner air by oval & round windows -2 passages for air -auditory or eustachian tube: opens into pharynx, equalizes pressure -passage to mastoid air cells in mastoid process -ossicles: malleus, incus, stapes: transmit vibrations from eardrum to oval window -oval window: connection btwn middle & inner ear. foot of the stapes rests here & is held in place by annular ligament hearing hearing relies on sensory hair cells in organs (the ear) that respond to the vibrations of sound waves sound waves -exist as variations of pressure in a medium such as air. -are created by the vibration of an object, which causes the air surrounding it to vibrate -the vibrating air then causes the human eardrum to vibrate, which the brain interprets as sound vibrations produce... soundwaves volume or loudness function of wave amplitude pitch function of wave frequency timbre resonance quality or overtones of sound eardrum tympanic membrane In the cochlea... vibrations transmitted from the eardrum through the fluid in the inner ear make the basilar membrane vibrate, bending the hair cells against the tectorial membrane and generating action potentials in afferent neurons that lead to auditory regions of the brain cochlea a spiraled, hollow, conical chamber of bone -structures include: scala vestibuli, scala tympani, and scala media scala vestibuli (containing perilymph), lies superior to the cochlear duct; abuts the oval window (is on outer side of cochlea) scala tympani (containing perilymph), lies inferior to the scala media; terminates at the round window (is on outer side of cochlea) scala media (containing endolymph), which is the membranous cochlear duct containing the organ of Corti Endolymph & perilymph both contain electrolytes & proteins Endolymph is rich in potassium (which produces an ionic, electrical potential) Perilymph is rich in Na+ Basilar Membrane -forms part of floor of cochlear duct -anchors sensory hair cells in Organ of Corti -vibrates in response to vibrations transmitted through inner ear -15,000 hair cells located along basilar membrane -synapse to afferent neurons -afferent neurons=auditory nerve -> auditory center in temporal lobe of brain Organ of Corti -located in the cochlear duct & contains sensory hair cells that detect the sound vibrations transmitted to the inner ear. -is the sensory organ of hearing, which is distributed along the partition separating fluid chambers in the coiled tapered tube of the cochlea -is the sensory epithelium, a cellular layer on the basilar membrane Detection of sound vibrations -hair cells arranged in rows. of inner hair cells (responsible for hearing) & outer hair cells (regulate tension on basilar membrane) -hair bundle: stereocilia of one inner hair cell -tip link (gating spring) attaches tip of each stereocilium in a hair bundle to the side of the next longer stereocilium. -as stereocilia bend, they open K+ gates (mechanically gated ion channel) Basilar membrane -narrow near the oval window; wider at the outer end of cochlear duct -high pitched sounds (which are comprised of high frequency vibrations) vibrate the basilar membrane at its narrow, beginning end -low pitched sounds vibrate the basilar membrane near the wider, farther end Basilar membrane 2 -hair cells in the organ of Corti are "tuned" to certain sound frequencies by way of their location in the cochlea due to the degree of stiffness in the basilar membrane -stiffness is due to the thickness & width of the basilar membrane -basilar membrane is stiffest & narrowest near its beginning (at the oval window, where the ossicle bones transmit the vibrations coming from the eardrum) -here, it allows only high-frequency vibrations to move the basilar membrane, and thus the hair cells Basilar membrane 3 -the farther from oval window, the less stiff the basilar membrane is, & thus the less sensitive to high frequencies -low frequencies travel down the tube, & the less-stiff membrane is moved most easily by them where the reduced stiffness allows -as the basilar membrane gets less & less stiff, it responds better to lower frequencies Sounds we hear -combinations of vibrations at different frequencies & intensities occurring simultaneously w/ different degrees of force along the basilar membrane of the cochlea -specific action potentials generated in specific neurons are transmitted to auditory centers in the brain, integrated, result in our perception of a specific sound (voice, motor, bird) Balance mechanoreceptors in humans: vestibular apparatus -3 semicircular canals -2 fluid filled chambers: utricle & saccule -perceives position & motion of head Semicircular canals filled w/ endolymph, detects rotational motion. oriented perpendicular Ampulla of a semicircular canal Ampulla (region at base of a semicircular canal) detects rotational movement of the head/body & causes movement of the endolymph which displaces the cupula & bends the sensory hair cell -> generates action potentials in afferent neurons that synapse w/ hair cells Utricle & Saccule (fluid filled chambers) -oriented 30 degrees to each other -info about head position (up/down) & changes in rate of linear motion of body -contain sensory hair cells w/ stereocilia -hair cells have a membrane that contains otoliths: calcium carbonate crystals -when head tilts or body moves non-linearly, otolithic membrane moves -> bends hair cells -> neurotransmitter release -> action potential -> brain perceives movement Balance: In sum -gelatinous mass moves in response to gravity, bending hair cells & initiating action potentials -otoliths stimulate hair cells w/ varying frequencies -patterns of stimulation translated by brain into specific info about head position or acceleration Please allow access to your computer’s microphone to use Voice Recording. 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Exploring the Potential of Voice User Interface (VUI) in Web Development Welcome to the dawn of a new era in web development, where interaction transcends the traditional keyboard and mouse. Imagine a world where your voice shapes the digital landscape—the potential of Voice User Interface (VUI) is an unfolding revolution. In this realm, websites and applications respond not just to your clicks but to your spoken commands, creating a seamless and intuitive user experience. Join us on a journey, with the skilled US Logo and Web, through the transformative landscape of VUI in web development, where innovation meets the spoken word, redefining how we engage with the digital universe. The fusion of artificial intelligence and voice recognition technology has birthed a paradigm where users communicate with technology in a natural, conversational manner. No longer confined to traditional input methods, Voice User Interface (VUI) introduces a dynamic dimension where your voice becomes the catalyst for seamless interactions, offering an immersive and hands-free experience. This innovative intersection of AI and voice technology is reshaping the digital landscape, paving the way for more intuitive, accessible, and engaging web development services that respond to the cadence of human speech. The Evolution of Interaction: AI and Voice Recognition The digital landscape undergoes a transformative shift with the evolution of interaction powered by artificial intelligence and voice recognition. This synergy introduces a new era where responsive web design is a necessity for US businesses, engaging users in a manner that mirrors natural, conversational communication. The intricate dance between AI algorithms and voice recognition systems marks a departure from traditional interaction methods, opening doors to a more dynamic and personalized user experience. Breaking the Constraints: Beyond Traditional Input Methods Voice User Interface (VUI) emerges as a liberator, breaking the constraints imposed by traditional input methods, much like choosing the right web development framework. No longer tethered to keyboards or touchscreens, users can seamlessly interact with technology using their voice, just as businesses can navigate the digital landscape with the optimal web development framework. This departure from the norm introduces a level of freedom and accessibility that transcends the limitations of conventional input mechanisms, making technology more inclusive and user-friendly. Voice User Interface (VUI): Unleashing a Dynamic Dimension Voice becomes the interface, unleashing a dynamic dimension in user interactions through Voice User Interface (VUI). This innovative approach transforms how users navigate and engage with websites and applications. The spoken word evolves into a powerful tool, steering the digital experience in response to users’ natural language, making interactions more intuitive and human-centric. Your Voice as a Catalyst: Transforming Interactions In the realm of VUI, your voice becomes the catalyst for transforming interactions. It goes beyond simple commands; it becomes a conduit for expressing intent, driving a profound shift in how users communicate with and command technology. This transformation elevates user engagement, making the digital experience more conversational, responsive, and tailored to individual preferences. Immersive Experiences: The Hands-Free Advantage of VUI VUI introduces a hands-free advantage, fostering immersive experiences where users can interact with technology without lifting a finger. This paradigm shift in user engagement eliminates the need for physical interaction devices, offering a seamless and unencumbered experience. The hands-free advantage not only enhances convenience but also opens doors to a range of applications where physical interaction was once a barrier. Innovation Intersection: AI and Voice Technology Unite At the intersection of innovation, artificial intelligence and voice technology unite to redefine the possibilities of digital interaction. The synergy of AI algorithms interpreting natural language and voice recognition understanding spoken commands creates a harmonious intersection where technology comprehends and responds to users in a manner mirroring human communication. This union marks a pivotal moment in the ongoing evolution of user interfaces. Reshaping the Digital Landscape: VUI in Action VUI in action reshapes the digital landscape by offering a transformative user experience. As voice commands seamlessly translate into digital actions, the traditional boundaries between users and technology blur. Websites and applications, once reliant on visual and tactile inputs, now respond dynamically to the vocal cues of users, creating an interactive environment that adapts to individual preferences. Paving the Way: More Intuitive and Accessible Web Development The advent of VUI paves the way for more intuitive and accessible web development. Designing interfaces that respond to voice commands requires a shift in the approach to user experience and accessibility. Developers are challenged to create interfaces that not only understand spoken language but also prioritize inclusivity, ensuring that VUI becomes a tool for all users, regardless of physical abilities or limitations. Engaging the Cadence: VUI Responding to Human Speech VUI takes user engagement to a new level by responding to the cadence and nuances of human speech. The rhythm, tone, and inflections in spoken language become integral components in user interactions. This responsiveness to the intricacies of human speech enhances the conversational nature of VUI, creating a more natural and enjoyable experience for users as they engage with digital platforms. Web Development Services Embrace the VUI Revolution As the VUI revolution unfolds, web development services embrace the shift towards voice-activated interfaces. The expertise of web development services becomes crucial in designing and implementing VUI seamlessly into websites and applications. From integrating voice recognition algorithms to optimizing user interfaces for voice commands, web development services play a pivotal role in ensuring a smooth transition and harnessing the full potential of this transformative technology. Unfolding the Era: Voice-Activated Digital Experiences VUI heralds the era of voice-activated digital experiences, where the spoken word becomes the primary driver of interactions. This unfolding era represents a departure from conventional modes of engagement, inviting users to navigate and control digital environments effortlessly through their voice. The possibilities are boundless as voice-activated digital experiences redefine the way we interact with technology, offering a glimpse into a future where the spoken word guides us through a seamlessly integrated and immersive digital landscape. Conclusion: In conclusion, the exploration of Voice User Interface (VUI) in web development unveils a revolutionary era where technology converges with the spoken word, reshaping user interactions and digital landscapes. This dynamic intersection of artificial intelligence and voice recognition technology propels us beyond traditional constraints, offering an immersive, hands-free, and intuitive digital experience. VUI’s transformative power is evident in its ability to respond to the cadence of human speech, breaking free from conventional input methods. As web development services embrace the VUI revolution, the era of voice-activated digital experiences unfolds, paving the way for more inclusive, accessible, and engaging interactions. This unfolding era represents not just a shift in technology but a redefinition of how users engage with the digital realm, propelling us into a future where the spoken word guides us through seamlessly integrated and immersive digital landscapes. FAQs: How does Voice User Interface (VUI) differ from traditional input methods in web development? VUI liberates users from keyboards and touchscreens, allowing seamless interaction through spoken commands, creating a more dynamic and inclusive digital experience. 2. What role does artificial intelligence play in the evolution of VUI in web development? Artificial intelligence powers VUI by interpreting natural language and enabling voice recognition, fostering a harmonious intersection that mirrors human communication in digital interactions. 3. How does VUI contribute to more intuitive and accessible web development? VUI challenges developers to design interfaces that respond to voice commands, prioritizing inclusivity and accessibility for users of all abilities, thereby paving the way for a more user-friendly digital environment. 4. Why is the responsiveness of VUI to the cadence of human speech significant? VUI’s responsiveness to the nuances of human speech enhances the conversational nature of interactions, creating a natural and enjoyable user experience in navigating digital platforms. 5. What is the role of web development services in the VUI revolution? Web development services are crucial in seamlessly implementing VUI into websites and applications, integrating voice recognition algorithms, and ensuring a smooth transition to harness the full potential of this transformative technology.
V593. Consider reviewing the expression of the 'A = B == C' kind. The expression is calculated as following: 'A = (B == C)'. V593. Consider reviewing the expression of the 'A = B == C' kind. The expression is calculated as following: 'A = (B == C)'. Анализатор обнаружил потенциальную ошибку в выражении, которое, скорее всего, работает не так, как задумывал программист. Данный тип ошибок чаще всего можно встретить в выражениях, где одновременно проверяется результат работы функции и выполняется присваивание. Рассмотрим простой пример: if (handle = Foo() != -1) Создавая такой код, программист, как правило, хочет выполнить действия в следующем порядке: if ((handle = Foo()) != -1) Но приоритет оператора '!=' выше, чем приоритет оператора '='. Поэтому выражение вычистится так: if (handle = (Foo() != -1)) Чтобы исправить ошибку, можно использовать скобки. Ещё лучше не жадничать на количестве строк кода. Текст вашей программы станет более читабелен, если написать так: handle = Foo(); if (handle != -1) Рассмотрим, как подобная ошибка может выглядеть на практике: if (hr = AVIFileGetStream(pfileSilence, &paviSilence, typeAUDIO, 0) != AVIERR_OK) { ErrMsg("Unable to load silence stream"); return hr; } Проверка в коде, что произошла ошибка, работает корректно и будет выдано сообщение "Unable to load silence stream". Беда в том, что переменная 'hr' будет хранить не код ошибки, а значение 1. Исправленный вариант кода: if ((hr = AVIFileGetStream(pfileSilence, &paviSilence, typeAUDIO, 0)) != AVIERR_OK) { ErrMsg("Unable to load silence stream"); return hr; } Анализатор не всегда выдает предупреждения, обнаруживая конструкцию вида "if (x = a == b)". Например, анализатор понимает, что данный код безопасен: char from; char to; bool result; ... if (result = from == to) {} Примечание. Если анализатор все-таки выдал ложное предупреждение, то есть два способа устранить его: 1) Добавить дополнительные скобки. Пример: "if (x = (a == b))". 2) Использовать комментарий для подавления предупреждения. Пример: "if (x = a == b) //-V593". Согласно Common Weakness Enumeration, потенциальные ошибки, найденные с помощью этой диагностики, классифицируются как CWE-783. Взгляните на примеры ошибок, обнаруженных с помощью диагностики V593. А ты совершаешь ошибки в коде? Проверь с помощью PVS-Studio Статический анализ кода для C, C++ и C# goto PVS-Studio; Мы используем cookie-файлы для анализа событий на нашем веб-сайте, что позволяет улучшить наш контент и сделать взаимодействие с пользователем более удобным. Продолжая просмотр страниц нашего веб-сайта, вы принимаете условия использования этих файлов. Узнайте подробнее о cookie-файлах и политике конфиденциальности или скройте это уведомление, нажав на кнопку. Подробнее → Не показывать
0 If I have a grid view like this and I want to add more columns. One column will display buttons and the other one - notes. enter image description here 1 • Did you have any additional questions that we didn't address in our responses? Commented May 26, 2016 at 19:47 3 Answers 3 2 You could have a "More details" button/link that opens a modal window with additional fields. mockup download bmml source – Wireframes created with Balsamiq Mockups Clicking on "More details" would open a popup similar to the one below. mockup download bmml source 3 • What about if i want to add more actions not just info Commented May 12, 2016 at 9:40 • 1 @AnynameDonotcare Modals can house all sorts of content: data, media, files, interactive tools, games, etc... You could have an "Actions" section in your modal that contains all available actions you can do with a particular piece of data. Commented May 12, 2016 at 13:25 • @AnynameDonotcare Maybe you could change the term "More details..." to say something else, but this is the way I would implement this. It effectively shows only the most important information/actions upfront, then hides the additional information/actions in the modal to avoid excessive clutter in the table. Commented May 12, 2016 at 14:46 2 What is shocking in your design is that 40% of the space is taken by action button and not by the main information. The buttons are taking to much space. Promote the content, not the tools. Some icons are very generic and don't need explanation. I don't speak Arabic but I can tell that the first button is "edit", second is "print", third "validate", fourth "settings" or "batch". Display only the icon and remove the text. Place the primary / most used icons first and directly accessible, place the rest into a more menu (like said maxathousand). 1 I have a suggestion, but it depends on how complex your build can be (i.e., what are the limitations of the build/developers). A while back, I actually spent some time building out table-design guidelines for College Board. One thing that became very useful was allowing "excess columns" to reside in an expanded portion of a row, displayed in a stacked view rather than as columns. The benefit of this is you can view multiple expanded views at a time (might get messy, but it's doable). It also allows for quicker reference between normal and excess information, and between the excess information of two or more rows. This also freed up modals to serve more as views for key actions (batch actions, printing, editing, etc.), as opposed to containers of information. Expandable rows in a table Your Answer By clicking “Post Your Answer”, you agree to our terms of service and acknowledge you have read our privacy policy. Not the answer you're looking for? Browse other questions tagged or ask your own question.
Nobelprize.org Nobel Prizes and Laureates Nobel Prizes and Laureates The Nobel Prize in Physics 1972 John Bardeen, Leon N. Cooper, Robert Schrieffer Share this: Award Ceremony Speech Presentation Speech by professor Stig Lundqvist, Chalmers University of Technology Translation from the Swedish text Your Royal Highnesses, Ladies and Gentlemen, The 1972 Nobel Prize for physics has been awarded to Drs John Bardeen, Leon N. Cooper and J. Robert Schrieffer for their theory of superconductivity, usually referred to as the BCS-theory. Superconductivity is a peculiar phenomenon occurring in many metallic materials. Metals in their normal state have a certain electrical resistance, the magnitude of which varies with temperature. When a metal is cooled its resistance is reduced. In many metallic materials it happens that the electrical resistance not only decreases but also suddenly disappears when a certain critical temperature is passed which is a characteristic property of the material. This phenomenon was discovered as early as 1911 by the Dutch physicist Kamerlingh Onnes, who was awarded the Nobel Prize for Physics in 1913 for his discoveries. The term superconductivity refers to the complete disappearance of the electrical resistance, which was later verified with an enormous accuracy. A lead ring carrying a current of several hundred ampères was kept cooled for a period of 2 1/2 years with no measurable change in the current. An important discovery was made in the thirties, when it was shown that an external magnetic field cannot penetrate a superconductor. If you place a permanent magnet in a bowl of superconducting material, the magnet will hover in the air above the bowl, literally floating on a cushion of its own magnetic field lines. This effect may be used as an example for the construction of friction-free bearings. Many of the properties of a metal change when it becomes superconducting and new effects appear which have no equivalent in the former's normal state. Numerous experiments have clearly shown that a fundamentally new state of the metal is involved. The transition to the superconductive state occurs at extremely low temperatures, characteristically only a few degrees above absolute zero. For this reason practical applications of the phenomenon have been rare in the past and superconductivity has been widely considered as a scientifically interesting but exclusive curiosity confined to the low temperature physics laboratories. This state of affairs is rapidly changing and the use of superconducting devices is rapidly increasing. Superconducting magnets are often used for example in particle accelerators. Superconductivity research has in recent years resulted in substantial advances in measuring techniques and an extensive used in the computer field is also highly probable. Advanced plans for the use of superconductivity in heavy engineering are also in existence. By way of an example, it may be mentioned that the transport of electric energy to the major cities of the world with the use of superconductive lines is being planned. Looking further ahead one can see, for example, the possibility of building ultrarapid trains that run on superconducting tracks. Superconductivity has been studied experimentally for more than sixty years. However, the central problem, the question of the physical mechanism responsible for the phenomenon remained a mystery until the late fifties. Many famous physicists tackled the problem with little success. The difficulties were related to the very special nature of the mechanism sought. In a normal metal the electrons more around individually at random, somewhat similar to the atoms in a gas, and the theory is, in principle, fairly simple. In superconductive metals the experiments suggested the existence of a collective state of the conduction electrons-a state in which the electrons are strongly coupled and their motion correlated so that there is a gigantic coherent state of macroscopic dimension containing an enormous number of electrons. The physical mechanism responsible for such a coupling remained unknown for a long time. An important step towards the solution was taken in 1950 when it was discovered simultaneously on theoretical and experimental grounds that superconductivity must be connected with the coupling of the electrons to the vibrations of the atoms in the crystal lattice. The conduction electrons are coupled to each other via these vibrations. Starting from this fundamental coupling of the electrons Bardeen, Cooper and Schrieffer developed their theory of superconductivity, published in 1957, which gave a complete theoretical explanation of the phenomenon of superconductivity. According to their theory the coupling of the electrons to the lattice oscillations leads to the formation of bound pairs of electrons. These pairs play a fundamental role in the theory. The complete picture of the mechanism of superconductivity appeared when Bardeen, Cooper and Schrieffer showed that the motion of the different pairs is very strongly correlated and that this leads to the formation of a gigantic coherent state in which a large number of electrons participate. It is this ordered motion of the electrons in the superconductive state in contrast to the random individual motion in a normal crystal that gives superconductivity its special properties. The theory developed by Bardeen, Cooper and Schrieffer together with extensions and refinements of the theory, which followed in the years after 1957, succeeded in explaining in considerable detail the properties of superconductors. The theory also predicted new effects and it stimulated intense activity in theoretical and experimental research which opened up new areas. These latter developments have led to new important discoveries which are being used in a number of interesting ways especially in the sphere of measuring techniques. Developments in the field of superconductivity during the last fifteen years have been greatly inspired by the fundamental theory of superconductivity and have strikingly verified the validity and great range of the concepts and ideas developed by Bardeens, Cooper and Schrieffer. Drs. Bardeen, Cooper and Schrieffer, You have in your fundamental work given a complete theoretical explanation of the phenomenon of superconductivity. Your theory has also predicted new effects and stimulated an intensive activity in theoretical and experimental research. The further developments in the field of superconductivity have in a striking way confirmed the great range and validity of the concepts and ideas in your fundamental paper from 1957. On behalf of the Royal Academy of Sciences I wish to convey to you the warmest congratulations and I now ask you to receive your prizes from the Hands of His Royal Highness the Crown Prince. From Nobel Lectures, Physics 1971-1980, Editor Stig Lundqvist, World Scientific Publishing Co., Singapore, 1992 Copyright © The Nobel Foundation 1972 Share this: To cite this page MLA style: "Nobelprize.org". Nobelprize.org. Nobel Media AB 2014. Web. 26 Jun 2016. <http://www.nobelprize.org/nobel_prizes/physics/laureates/1972/presentation-speech.html> Recommended:
Can you calibrate a digital scale? Calibrating your digital scale is the best way to prolong the scale’s quality and overall lifetime. It only takes a few minutes, and when done correctly, guarantees the measurements you are taking are accurate each and every time. >> Click to read more << Consequently, how do I know if my scale is accurate? Weigh two objects together. 1. Place one object on the scale. Note the weight. Take it off and let the scale even back out. … 2. If it matches, the scale is accurate. If it doesn’t, try it again and see if it is off by the same number. If it is, it might be that your scale is always off by that amount. Just so, how do I reset my digital scale calibration? 1. Reset unit by pressing ON/OFF button and wait until “0.0” is shown. 2. Press and hold UNITS button for 3 seconds until “CAL” is displayed. Keeping this in consideration, how do you balance a weight scale? How do you calibrate a digital scale at home? How to Calibrate Digital Scales: A Step-by-Step Guide 1. Step 1: Gather Your Weights. … 2. Step 2: Turn on Your Scale. … 3. Step 3: Locate and Press Your Scale’s Calibration Button. … 4. Step 4 Put Your Weights on the Scale Platform. … 5. Step 5: Press the Calibration Button Again. … 6. Step 6: Reset the Scale. … 7. Step 7: Make Sure the Calibration Worked. How do you calibrate a digital scale without weights? First, turn the power on, then press the “Zero” or “Tare” button if there is one present. Wait patiently while the scale clears any remaining data from previous uses. It may take a second, but your scale should show a “0.00” weight once it is zeroed. How do you calibrate a scale with household items? Place a calibration weight, a U.S. coin, or household item on your scale. 1. Pennies made after 1983 weigh exactly 2.5 grams (0.088 oz). 2. Nickels made after 1866 weigh 5 grams (0.18 oz) 3. Dimes made after 1965 weigh 2.27 grams (0.080 oz) 4. Quarters made after 1965 weigh 5.67 grams (0.200 oz) How do you calibrate a weighing scale device? To calibrate a digital scale, set it on a flat surface and choose a weight. Turn the scale on and go into calibration mode. Put the weight on the scale and wait for it to calculate the total weight. If the weight is off, hit the calibration button and wait for it to recalibrate. How do you calibrate a weight max digital scale? Press and hold the calibrate key, which is marked “CAL.” Wait until “CAL” is displayed on the LCD screen. The calibration display will then read the zero point, “0.0.” Press the “CAL” key again and hold it for two to three seconds to wait for the scale to calibrate the zero point and display the full capacity. How do you use the soehnle kitchen scale? Is it necessary to calibrate a weighing scale? Balance or scale calibration is essential to achieve accurate weighing results. Ignoring this important service activity turns measuring into guesswork. In other words, it is negligent to weigh with a non-calibrated balance or scale. The accuracy of balances and scales becomes less reliable over time. What can I use as a 200g weight? Many items can amount to 200 grams, such as a roll of nickels and a cup of granulated white sugar. Other items of that weight include three C-cell batteries and an adult hamster. However, there are plenty of other everyday things that all weigh the same amount. What can I use for 500g calibration weight? • A Chinchilla. A popular pet, chinchillas are known to be friendly and soft. … • A Loaf of Bread. Most loaves of bread weigh about 500 grams. … • A Package of Beef. Meats such as beef are often sold by the pound or by the gram. … • A Bottle of Salad Dressing. … • Two Skeins of Yarn. … • 10 Medium Eggs. … • 100 Nickels. … • A Soccer Ball. Where are soehnle scales made? Germany Leave a Comment
20 votes 14answers 22k views What programming languages are good for novice testers to learn? I'm looking for a programming language to recommend to a friend considering teaching herself software testing. I have a friend who is interested in learning software testing, and I believe strongly ... 5 votes 3answers 336 views SQL “sandbox” for new QA staff training: DB structure, approach, deployment and maintenance? Complex DB solutions are not rare nowadays, so I have an idea to deploy a SQL "sandbox" for staff training before real projects come. Mainly I interested in 2 things to have: Oracle 11+ DB instance. ... 4 votes 3answers 90 views Fostering teamwork in a distributed organization What techniques have you used successfully to promote teamwork between developers and testers who are geographically dispersed? Here is some context. This morning I talked with someone who works for ... 4 votes 3answers 1k views Does the latest version of QTP have something similar to Selenium Grid? 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It might be VERY different - below are ... 1 vote 1answer 84 views Issue tracker that supports geo location Is there a free issue tracker software that permits to attach a location to an issue (preferably by marking the location on a map), permits to filter issues by location (preferably by displaying all ... 2 votes 1answer 816 views WebDriver Ruby binding: How to specify Firefox binary path in code? I'm writing my test cases in Windows7 with Ruby (v1.9.3-p194). Please, note that I want to specify Firefox binary path when using RemoteWebDriver, not FirefoxDriver. Here is the command line I used ... 6 votes 2answers 132 views Why should I have a “closed beta”? I am running a test on a new Rails app and am doing an open beta from the start. You see these companies with private betas, and I'm wondering, what is the exact point of a private beta. If you inform ... 3 votes 1answer 7k views Selenium - page load timeout Is it possible to set timeout for page load in Selenium test? 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Although, common perception is that dynamic analysis is testing itself, most of advanced sources (but AFAIK also the ISTQB) distinct between: dynamic testing ... 0 votes 4answers 277 views Automated network capture and analysis? I was curious to know if anyone has done any work on automated network analysis such as starting network traffic capture (to file) before executing some test, perform some test, then stop capture, ... 6 votes 5answers 5k views Is there a guide or systematic way of writing test scenarios? Are there best practices in coming up with effective test scenarios? Are there documented test scenarios that can be reused across different systems? 3 votes 3answers 1k views Upgrade testing strategy diagram Does anyone know of any diagram or tool that can help me think and chalk out a strategy for upgrade testing? Suppose there is a server side component that can have add-ons, plugins if you will. 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I tried ... 1 vote 2answers 191 views Selenium Selenium.Core v1.1.4322 - XPath not working I've got the following XPath expression's working in firebug, that returns the correct element(s): $x("(//input[@class='intervalInput timeInput'])") returns two elements. ... 1 vote 2answers 579 views Not able to identify tab in Selenium Below is code for tab - account.This is third tab. I have to go on tab but I am not able to identify <td class="buttons" nowrap="nowrap" title="Shows data of bank account" ... 1 vote 4answers 367 views Performance testing resources Please share any good performance testing resources which you have come across in past or referred to while working or learning about performance testing. 13 votes 11answers 912 views What are the forms of testing that testers commonly forget? Today I re-realized the importance of sharing a screen with your users to see exactly what they're doing and the paths they are taking through your software. 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The settings for automating Vaddin application ... -8 votes 1answer 887 views Pre-requisites & Procedure to install selenium tool on Windows 7 Operating system? [closed] I would like to Install Selenium open source tool in my pc ( windows 7 OS) hence would like to know the procedure as well as pre-requisities . Thank you regards venkat 1 vote 0answers 156 views Strategies for testing .ics or .ical file compatibility with iOS devices I need to test .ics and .ical files for compatibility with the native calendar app on iOS devices. If I purchase an iOS dev kit will this allow me to use the native calender application under ... 1 vote 0answers 418 views Simple SVN post-commit hook for Jenkins on Windows Server 2008 [closed] I have VisualSVN Server holding several repos with PHP applications on Windows Server 2008 machine. 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[closed] I created an Excel file name "readdata.xls" and entered some inputs like UserName Password Akshit Ak12345 Ashwika AS12345 I've written a code like this .... package ... 4 votes 2answers 810 views Selenium Server and Windows Server compatibility I want to run Selenium Server on a recent version of Windows Server so as to be able to test IE browsers as well as Firefox, Chrome, etc. My choices for Windows Server are 2008 R2, 2008, or 2003. ... -5 votes 1answer 2k views ArrayOutOfBoundException when reading test data from Excel [closed] I am very new to Selenium RC. Please, help in my question. I installed jxl.jar file (Java library for Excel) and added it to Eclipse build path. I created an Excel file as readdata.xls where I ... 9 votes 3answers 569 views How to create a 'quality-centric culture' within a team/organization? I have recently talked to the head of the quality department of another company and they struggle with the problem that the developers still think quality is actually the QA's job. They work in ... 2 votes 1answer 519 views Telecom testing - any tool integration and use of test frameworks? Telecom testing is an interesting niche area, and you either use expensive commercial tool solutions, or open source solutions to do testing (e.g. SIPp), but they're not always one-size-fits-all. I ... 7 votes 3answers 4k views How to finish the testing in the same iteration/sprint? I am aware of the fact that agile testing is an ongoing process and should happen in parallel to development. But how is that actually achieved? Is it even possible to fix all found issues in the same ... 4 votes 2answers 4k views Web application GUI automation best practices Our team develops automation solutions for various web based products. 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Hibernate - Examples Advertisements Let us now take an example to understand how we can use Hibernate to provide Java persistence in a standalone application. We will go through the different steps involved in creating a Java Application using Hibernate technology. Create POJO Classes The first step in creating an application is to build the Java POJO class or classes, depending on the application that will be persisted to the database. Let us consider our Employee class with getXXX and setXXX methods to make it JavaBeans compliant class. A POJO (Plain Old Java Object) is a Java object that doesn't extend or implement some specialized classes and interfaces respectively required by the EJB framework. All normal Java objects are POJO. When you design a class to be persisted by Hibernate, it is important to provide JavaBeans compliant code as well as one attribute, which would work as index like id attribute in the Employee class. public class Employee { private int id; private String firstName; private String lastName; private int salary; public Employee() {} public Employee(String fname, String lname, int salary) { this.firstName = fname; this.lastName = lname; this.salary = salary; } public int getId() { return id; } public void setId( int id ) { this.id = id; } public String getFirstName() { return firstName; } public void setFirstName( String first_name ) { this.firstName = first_name; } public String getLastName() { return lastName; } public void setLastName( String last_name ) { this.lastName = last_name; } public int getSalary() { return salary; } public void setSalary( int salary ) { this.salary = salary; } } Create Database Tables Second step would be creating tables in your database. There would be one table corresponding to each object, you are willing to provide persistence. Consider above objects need to be stored and retrieved into the following RDBMS table − create table EMPLOYEE ( id INT NOT NULL auto_increment, first_name VARCHAR(20) default NULL, last_name VARCHAR(20) default NULL, salary INT default NULL, PRIMARY KEY (id) ); Create Mapping Configuration File This step is to create a mapping file that instructs Hibernate how to map the defined class or classes to the database tables. <?xml version = "1.0" encoding = "utf-8"?> <!DOCTYPE hibernate-mapping PUBLIC "-//Hibernate/Hibernate Mapping DTD//EN" "http://www.hibernate.org/dtd/hibernate-mapping-3.0.dtd"> <hibernate-mapping> <class name = "Employee" table = "EMPLOYEE"> <meta attribute = "class-description"> This class contains the employee detail. </meta> <id name = "id" type = "int" column = "id"> <generator class="native"/> </id> <property name = "firstName" column = "first_name" type = "string"/> <property name = "lastName" column = "last_name" type = "string"/> <property name = "salary" column = "salary" type = "int"/> </class> </hibernate-mapping> You should save the mapping document in a file with the format <classname>.hbm.xml. We saved our mapping document in the file Employee.hbm.xml. Let us see little detail about the mapping document − • The mapping document is an XML document having <hibernate-mapping> as the root element which contains all the <class> elements. • The <class> elements are used to define specific mappings from a Java classes to the database tables. The Java class name is specified using the name attribute of the class element and the database table name is specified using the table attribute. • The <meta> element is optional element and can be used to create the class description. • The <id> element maps the unique ID attribute in class to the primary key of the database table. The name attribute of the id element refers to the property in the class and the column attribute refers to the column in the database table. The type attribute holds the hibernate mapping type, this mapping types will convert from Java to SQL data type. • The <generator> element within the id element is used to generate the primary key values automatically. The class attribute of the generator element is set to native to let hibernate pick up either identity, sequence or hilo algorithm to create primary key depending upon the capabilities of the underlying database. • The <property> element is used to map a Java class property to a column in the database table. The name attribute of the element refers to the property in the class and the column attribute refers to the column in the database table. The type attribute holds the hibernate mapping type, this mapping types will convert from Java to SQL data type. There are other attributes and elements available, which will be used in a mapping document and I would try to cover as many as possible while discussing other Hibernate related topics. Create Application Class Finally, we will create our application class with the main() method to run the application. We will use this application to save few Employee's records and then we will apply CRUD operations on those records. import java.util.List; import java.util.Date; import java.util.Iterator; import org.hibernate.HibernateException; import org.hibernate.Session; import org.hibernate.Transaction; import org.hibernate.SessionFactory; import org.hibernate.cfg.Configuration; public class ManageEmployee { private static SessionFactory factory; public static void main(String[] args) { try { factory = new Configuration().configure().buildSessionFactory(); } catch (Throwable ex) { System.err.println("Failed to create sessionFactory object." + ex); throw new ExceptionInInitializerError(ex); } ManageEmployee ME = new ManageEmployee(); /* Add few employee records in database / Integer empID1 = ME.addEmployee("Zara", "Ali", 1000); Integer empID2 = ME.addEmployee("Daisy", "Das", 5000); Integer empID3 = ME.addEmployee("John", "Paul", 10000); / List down all the employees / ME.listEmployees(); / Update employee's records / ME.updateEmployee(empID1, 5000); / Delete an employee from the database / ME.deleteEmployee(empID2); / List down new list of the employees / ME.listEmployees(); } / Method to CREATE an employee in the database / public Integer addEmployee(String fname, String lname, int salary){ Session session = factory.openSession(); Transaction tx = null; Integer employeeID = null; try { tx = session.beginTransaction(); Employee employee = new Employee(fname, lname, salary); employeeID = (Integer) session.save(employee); tx.commit(); } catch (HibernateException e) { if (tx!=null) tx.rollback(); e.printStackTrace(); } finally { session.close(); } return employeeID; } / Method to READ all the employees / public void listEmployees( ){ Session session = factory.openSession(); Transaction tx = null; try { tx = session.beginTransaction(); List employees = session.createQuery("FROM Employee").list(); for (Iterator iterator = employees.iterator(); iterator.hasNext();){ Employee employee = (Employee) iterator.next(); System.out.print("First Name: " + employee.getFirstName()); System.out.print(" Last Name: " + employee.getLastName()); System.out.println(" Salary: " + employee.getSalary()); } tx.commit(); } catch (HibernateException e) { if (tx!=null) tx.rollback(); e.printStackTrace(); } finally { session.close(); } } / Method to UPDATE salary for an employee / public void updateEmployee(Integer EmployeeID, int salary ){ Session session = factory.openSession(); Transaction tx = null; try { tx = session.beginTransaction(); Employee employee = (Employee)session.get(Employee.class, EmployeeID); employee.setSalary( salary ); session.update(employee); tx.commit(); } catch (HibernateException e) { if (tx!=null) tx.rollback(); e.printStackTrace(); } finally { session.close(); } } / Method to DELETE an employee from the records / public void deleteEmployee(Integer EmployeeID){ Session session = factory.openSession(); Transaction tx = null; try { tx = session.beginTransaction(); Employee employee = (Employee)session.get(Employee.class, EmployeeID); session.delete(employee); tx.commit(); } catch (HibernateException e) { if (tx!=null) tx.rollback(); e.printStackTrace(); } finally { session.close(); } } } Compilation and Execution Here are the steps to compile and run the above mentioned application. Make sure, you have set PATH and CLASSPATH appropriately before proceeding for the compilation and execution. • Create hibernate.cfg.xml configuration file as explained in configuration chapter. • Create Employee.hbm.xml mapping file as shown above. • Create Employee.java source file as shown above and compile it. • Create ManageEmployee.java source file as shown above and compile it. • Execute ManageEmployee binary to run the program. You would get the following result, and records would be created in the EMPLOYEE table. $java ManageEmployee .......VARIOUS LOG MESSAGES WILL DISPLAY HERE........ First Name: Zara Last Name: Ali Salary: 1000 First Name: Daisy Last Name: Das Salary: 5000 First Name: John Last Name: Paul Salary: 10000 First Name: Zara Last Name: Ali Salary: 5000 First Name: John Last Name: Paul Salary: 10000 If you check your EMPLOYEE table, it should have the following records − mysql> select from EMPLOYEE; +----+------------+-----------+--------+ \| id \| first_name \| last_name \| salary \| +----+------------+-----------+--------+ \| 29 \| Zara \| Ali \| 5000 \| \| 31 \| John \| Paul \| 10000 \| +----+------------+-----------+--------+ 2 rows in set (0.00 sec mysql> Advertisements E-Books Store
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JQuery loading of Google Charts I frequently work with Google Charts, and got tired of hand coding each of the charts individually, so I wrote a JQuery script (which I may convert into a plugin) that detects the data dumped directly into the data element of a div assigned to the chart. I've posted the code to JSFiddle. Feel free to use and extend this as needed. This is convenient, as you don't need to build a bunch of Javascript data to implement the chart. Additionally, you could convert this to pull data from a JSON source (since the data is in JSON format).
It has been proven time and time again that worrying too much and for too long can manifest itself as physical sickness. With the current state of US politics, it is no surprise that there are some people who are worrying more than ever. Every day they are greeted with the news of what the President has said this time, and the destruction and negative aftermath that followed. There is a lot going on and a lot of tragedies happening, so it is not shocking that a lot of the nation is getting very worried about the future. Unfortunately, this may be causing more sick days than anyone realizes. To worry is a form of stress, and it can have multiple negative effects on your health, especially if you never give yourself a break from it. If you are a constant worrier, you can cause yourself some real emotional damage that will also eventually come with some serious physical effects as well. Worrying causes stress hormones, and is the accumulation of these that will start producing negative things in your body. Your glands, nervous system and your heart can all be afflicted and this can end up causing heart disease, ulcers, and increase your risk of a heart attack. Other less serious symptoms can include back pain, constipation, headaches and muscle tension. You will also open yourself and become more susceptible to infectious diseases as the constant worry weakens your immune system. Worrying may also disturb your peace of mind, it can cause lack of sleep, cause you to isolate yourself socially and even coax you into a depression. Worry differs from fear in that when you fear something, there is a concrete obstacle in your way. Excessive worry can make everything feel like a threat, and cause anxiety and panic attacks to come on. There is no denying the effect that excessive worry can have on our minds as well as our bodies, so it is in your best interest to stop worrying so much about what is going on in Washington. The politics might literally be making you sick, but there are ways to avoid this, if you know what the real problem is. It is also winter time and there is not a lot of sunshine available; this does not help the situation. A lack of sunshine may also contribute to increased illness frequency. One of the first things it can do to some, is pull them into a depression as things around them start looking and feeling dreary. Depression itself can cause a whole list of physical problems. Something that might be beneficial is to get a massage. Put your mind and body fully at ease, and become more positive and proactive about your thoughts. If you constantly think negativity, that is what will manifest in your life. Alternatively, if you think good and positive thoughts, this is what you will begin to experience. There are a few other solutions you can try – one being more exercise. Try to get out there and move your body a little bit, without watching the news. Don’t let your mind be bogged down by how dark it is outside and what is happening in the world. Following a healthier diet will make you feel better,as well as provide extra energy. Leafy greens, fruits, and vegetables are good places to start when trying to add good things to your diet. Drinking more water is also a good idea. Most people tend to think they are drinking enough water but they aren’t – our bodies need more than we sometimes think! Lastly, try to improve and regulate your sleep schedule. Our bodies heal when we sleep, so give your body the time it needs to heal you. Finally if you feel that your worry is having serious effects on your lifestyle, contact your mental health counselor or your doctor as there may be clinical issues that need addressing. The President will more than likely to continue to say things that may make us concerned but you can guared your mind and body for a healthier you. CategoryHealth, Wellness Write a comment: * Your email address will not be published. © 2018 6th Sense Health & Wellness Winston Salem, NC \| Specialist in Massage, Acupuncture and other wellness services \| Medical Disclaimer\| logo-footer STAY CONNECTED WITH US:
Journal of Biomedical Imaging and Bioengineering All submissions of the EM system will be redirected to Online Manuscript Submission System. Authors are requested to submit articles directly to Online Manuscript Submission System of respective journal. Reach Us +1 (202) 780-3397 Commentary - Journal of Biomedical Imaging and Bioengineering (2022) Volume 6, Issue 1 Wavelength maps for X-ray computed tomography frameworks. Zheng Wang* Department of Gastroenterology, Xiangya Hospital, Central South University, Changsha, China *Corresponding Author: Zheng Wang Department of Gastroenterology Xiangya Hospital, Central South University Changsha, China E-mail: liuxwe11@csu.edu.cn Received: 31-Dec-2021, Manuscript No. AABIB-22-54572; Editor assigned: 03-Jan-2022, Pre QC No. AABIB -22-54572 (PQ); Reviewed: 17-Jan-2022, QC No. AABIB-22-54572; Revised: 22-Jan-2022; AABIB-22-54572 (R); Published: 29-Jan-2022, DOI: 10.35841/aabib-6.1.101 Citation: Yideng Jiang. Wavelength maps for X-ray computed tomography frameworks. J Biomed Imag Bioeng. 2022;4(1):101. Visit for more related articles at Journal of Biomedical Imaging and Bioengineering Abstract Amplitude–wavelength (AW) maps or “Stedman diagrams” are frequently utilized to supply a graphical representation of the impediments and capabilities of surface measuring disobedient. This paper presents an approach for setting the parameter limitations of X-ray computed tomography (CT) in terms of determination and measuring extend for the reason of speaking to the execution of mechanical CT frameworks on an AW outline. Such AW outline will permit instrument clients to rapidly compare the CT instrument execution to other measuring frameworks. Cases of the development of AW maps for distinctive working capabilities of X-ray CT frameworks, and based on test information, are given. Polypropylene, aluminium, and steel are three work piece materials considered for deciding a few of the confinements of measuring capability for the maps created in this paper Keywords Computed tomography, Non-destructive inspection X-ray, CT Measurement X-ray, Pillar constriction Introduction Radiography's roots and fluoroscopy's beginnings can both be followed to 8 November 1895, when German material science teacher Wilhelm Conrad Rontgen found the X-ray and famous that, whereas it seem pass through human tissue, it seem not pass through bone or metal. Rontgen alluded to the radiation as "X", to demonstrate that it was an obscure sort of radiation. He has gotten the primary Nobel Prize in Material science for his disclosure [1]. There are clashing accounts of his revelation since Röntgen had his lab notes burned after his passing, but typically a likely reproduction by his biographers. Rontgen was examining cathode beams employing a fluorescent screen painted with barium platinocyanide and a Crookes tube which he had wrapped in dark cardboard to shield its fluorescent gleam. He has taken note a swoon green gleam from the screen, almost 1 meter absent. Rontgen realized a few invisible rays coming from the tube were passing through the cardboard to form the screen gleam: they were passing through a dark question to influence the film behind itp [2]. The creation of pictures by uncovering a protest to X-rays or other high-energy shapes of electromagnetic radiation and capturing the coming about leftover bar (or "shadow") as an inactive picture is known as "projection radiography." The "shadow" may be changed over to light employing a fluorescent screen, which is at that point captured on photographic film, it may be captured by a phosphor screen to be "examined" afterward by a laser (CR), or it may straightforwardly actuate a lattice of solid-state detectors (DR—similar to a really expansive form of a CCD in an advanced camera). Bone and a few organs (such as lungs) particularly loan themselves to projection radiography [3]. DEXA, or bone densitometry, is utilized fundamentally for osteoporosis tests. It isn't projection radiography, as the X-rays are radiated in 2 contract bars that are filtered over the persistent, 90 degrees from each other. Ordinarily the hip (head of the femur), lower back (lumbar spine), or heel (calcaneum) are imaged, and the bone thickness (sum of calcium) is decided and given a number (a T-score). It isn't utilized for bone imaging, as the image quality isn't great sufficient to form an exact demonstrative picture for fractures, inflammation, etc. It can moreover be utilized to degree add up to body fat, in spite of the fact that typically not common [4]. Mechanical radiography could be a strategy of non-destructive testing where numerous sorts of made components can be inspected to confirm the inside structure and judgment of the example [5]. Mechanical Radiography can be performed utilizing either X-rays or gamma beams. Both are shapes of electromagnetic radiation. The distinction between different shapes of electromagnetic vitality is related to the wavelength. X and gamma beams have the most limited wavelength and this property leads to the capacity to enter, travel through, and exit different materials such as carbon steel and other metals. Particular strategies incorporate mechanical computed tomography. References 1. Jang J, Jung SE, Jeong WK, et al. Radiation doses of various CT protocols: a multicenter longitudinal observation study. J Korean Med Sci. 2016;3:24-31. 2. Indexed at, Google Scholar, Cross Ref 3. Schueler BA. The AAPM/RSNA physics tutorial for residents general overview of fluoroscopic imaging. Radiographics. 2000;20:1115-26. 4. Indexed at, Google Scholar, Cross Ref 5. Chalkley M, Listl S. First do no harm–The impact of financial incentives on dental X-rays. J Health Econ. 2018;58:1-9. 6. Indexed at, Google Scholar, Cross Ref 7. Barry K, Kumar S, Linke R. A clinical audit of anatomical side marker use in a paediatric medical imaging department. J Med Radiat Sci. 2016;63:148-54. 8. Indexed at, Google Scholar, Cross Ref 9. Seibert JA. Flat-panel detectors: how much better are they? Pediatr Radiol. 2006;36:173-81. 10. Indexed at, Google Scholar, Cross Ref Get the App
Open Access Expression of autophagy-related protein beclin-1 in malignant canine mammary tumors Contributed equally BMC Veterinary Research20139:75 https://doi.org/10.1186/1746-6148-9-75 Received: 18 December 2012 Accepted: 10 April 2013 Published: 11 April 2013 Abstract Background Autophagy is a self-catabolic mechanism that degrades unnecessary cellular components through lysosomal enzymes. Beclin-1, an autophagy-related protein, establishes the first connection between autophagy and tumorigenesis. The purpose of this study is to assess the Beclin-1 expression pattern and to determine its prognostic significance in patients with malignant canine mammary tumor (CMT). Results We examined Beclin-1 expression in 70 cases of malignant CMTs by immunohistochemistry. Cytoplasmic Beclin-1 expression was significantly weaker in cancer cells than in nearby normal mammary glands (p < 0.001). Low cytoplasmic expression (57.14%) was associated with older age, lower degree of tubular formation, increased mitotic activity, higher histologic grade, and extensive necrosis. Low nuclear expression (40%) was connected with older age, lower degree of tubular formation, extensive necrosis, and negative for Her2/neu overexpression. Univariate survival analysis showed that Beclin-1 cytoplasmic expression was a poor prognostic factor for overall survival rate (p < 0.001). Multivariate survival analysis demonstrated that Beclin-1 cytoplasmic expression is an independent prognostic factor (p = 0.016). Conclusions Loss of Beclin-1 is associated with aggressive clinicopathologic features and poor overall survival. The results suggest that Beclin-1 plays an important role in tumor progression of malignant CMTs. Keywords Autophagy Beclin-1 Canine mammary tumor Immunohistochemistry Background Canine mammary tumors (CMTs) are the most common neoplasms in intact female dogs. Approximately half of CMTs are malignant. Histologically, the majority of malignant CMTs are carcinomas, whereas approximately 10% are sarcomas. The spontaneously occurring malignant CMTs share many clinicopathologic and molecular characteristics with human breast cancers. The comparative analysis of human and dog genomes demonstrates the similarity of orthologous genes between the 2 species [1, 2]. Therefore, malignant CMTs can be used as a suitable animal model for oncogenesis research and treatment protocols. Programmed cell death is a genetically mediated process via internal or external signal pathways. Two types of programmed cell death, apoptosis and autophagy, have been subjects of increasing attention to scientists. Apoptosis involves the activation of catabolic enzymes in the signaling transduction pathway that leads to self-destruction. The term “autophagy” was first introduced in 1963 by de Duve, the discoverer of lysosomes [3]. Autophagy is a self-catabolic process that involves the degradation of intracellular structures and organelles by lysosomal enzymes [4]. Autophagy is essential for development, homeostasis, and survival, especially for stress adaption in an energy-deficient environment. It is also closely related to many pathologic processes, such as infections, metabolic disorders, neurodegeneration, and tumorigenesis [5]. Autophagy is regulated by a group of evolutionarily conserved genes, which were first discovered in yeast [6]. To date, more than 30 autophagy-related genes have been identified. The BECN1 gene is the mammalian orthologue of the yeast apg6/vps30, and was the first gene to establish a connection between autophagy and tumorigenesis [7]. Two research groups have shown that BECN1 heterozygous-deficient mice have a higher frequency of spontaneous tumors, whereas homozygous-deficient mice died early in embryogenesis because of defects in proamniotic canal closure [8, 9]. They concluded that BECN1 is a haplo-insufficient tumor suppressor gene. The Beclin-1 protein, which is encoded by the BECN1 gene, is involved in the signaling pathway of autophagy and is required for the nucleation of the phagophore and maturation of the autolysosome. Beclin-1 expression can indicate autophagic activity in cells. Beclin-1 expression and its association with clinicopathologic features have not been described in canine cancer. The aims of the study were to compare Beclin-1 expression patterns in normal mammary glands and malignant CMTs, to investigate the clinicopathologic significance of Beclin-1 expression, and to evaluate its impact on clinical outcomes. Results Patient characteristics This study comprised 70 cases of malignant CMTs, including 54 simple carcinomas, 11 complex carcinomas, and 5 sarcomas. The mean age of 69 dogs at the time of surgery was 11.3 ± 2.7 years (ranging from 4 to 18 years). The age of the remaining dog was unknown. In total, 16 of 70 (22.9%) dogs received ovario-hysterectomy prior to the surgical removal of tumors. The mean maximum tumor size was 4.3 ± 3.1 cm (ranging from 0.4 to 15.0 cm). The other clinicopathologic features, including tumor location, tubular formation, nuclear pleomorphism, mitotic count, histologic grade, lymphovascular invasion, necrosis, expressions of estrogen receptor and Her2, were summarized in Table 1. Table 1 Association of Beclin-1 expression pattern and clinicopathologic variables in 70 cases of malignant CMTs Beclin-1 cytoplasmic expression p value Beclin-1 nuclear expression p value Variable No. of cases Low High Low High Agea 11 years 36 15 (38.5%) 21 (70.0%) 0.009* 10 (37.0%) 26 (61.9%) 0.044* > 11 years 33 24 (61.5%) 9 (30.0%) 17 (63.0%) 16 (38.1%) Location of affected gland Cranial gland 24 14 (35.0%) 10 (33.3%) 0.953 9 (32.1%) 15 (35.7%) 0.890 Caudal gland 42 24 (60.0%) 18 (60.0%) 17 (60.7%) 25 (59.5%) Both 4 2 (5.0%) 2 (6.7%) 2 (7.1%) 2 (4.8%) Tumor size 3 cm 27 13 (32.5%) 14 (46.7%) 0.228 9 (32.1%) 18 (42.9%) 0.367 > 3 cm 43 27 (67.5%) 16 (53.3%) 19 (67.9%) 24 (57.1%) Histologic classification Simple carcinoma 54 30 (75.0%) 24 (80.0%) 0.877 23 (82.1%) 31 (73.8%) 0.640 Complex carcinoma 11 7 (17.5%) 4 (13.3%) 3 (10.7%) 8 (19.0%) Sarcoma 5 3 (7.5%) 2 (6.7%) 2 (7.1%) 3 (7.1%) Tubular formation > 10% of the tumor 45 19 (47.5%) 26 (86.7%) 0.001* 14 (50.0%) 31 (73.8%) 0.042* 10% of the tumor 25 21 (52.5%) 4 (13.3%) 14 (60.0%) 11 (26.2%) Nuclear pleomorphism Mild to moderate 46 17 (42.5%) 19 (63.3%) 0.084 12 (42.9%) 24 (57.1%) 0.241 Marked 34 23 (57.5%) 11 (36.7%) 16 (57.1%) 18 (42.9%) Mitotic count 10/10 HPFs 52 25 (62.5%) 27 (90.0%) 0.009* 18 (64.3%) 34 (81.0%) 0.118 >10/10 HPFs 18 15 (37.5%) 3 (10%) 10 (35.7%) 8 (19.0%) Histologic grade Grades 1 and 2 56 27 (67.5%) 29 (96.7%) 0.003* 18 (64.3%) 38 (90.5%) 0.007* Grade 3 14 13 (32.5%) 1 (3.3%) 10 (35.7%) 4 (9.5%) Lymphovascular invasion Absent 51 26 (65.0%) 25 (83.3%) 0.088 19 (67.9%) 32 (76.2%) 0.442 Present 19 14 (35.0%) 5 (16.7%) 9 (32.1%) 10 (23.8%) Necrosis Limited/no necrosis 41 17 (42.5%) 24 (80%) 0.002* 12 (42.9%) 29 (69.0%) 0.029* Extensive necrosis 29 23 (57.5%) 6 (20%) 16 (57.1%) 13 (31.0%) Estrogen receptor Negative 25 16 (40%) 9 (30%) 0.388 9 (32.1%) 16 (38.1%) 0.611 Positive 45 24 (60%) 21 (70%) 19 (67.9%) 26 (61.9%) Her2 overexpression Negative 51 29 (72.5%) 22 (73.3%) 0.938 24 (85.7%) 27 (64.3%) 0.048* Positive 19 11 (27.5%) 8 (26.7%) 4 (14.3%) 15 (35.7%) HPF, High power field. aThe age of one case is unknown. p < 0.05. Comparison of Beclin-1 expression in normal mammary glands and cancer cells The normal mammary glands near the cancer cells showed weak or moderate cytoplasmic reactivity and variable nuclear expression of Beclin-1 (Figure 1). The cancer cells displayed negative, weak, or moderate cytoplasmic staining, and ranged from non-reactivity to strong positivity of nuclear expression (Figure 2). The cytoplasmic Q score of normal mammary glands was significantly higher than that of cancer cells (p < 0.001). The difference of nuclear Q score between normal glands and cancer cells was not statistically significant (p = 0.130) (Figure 3). Figure 1 Beclin-1 expression in normal mammary glands of dogs. The normal mammary glands show diffuse moderate cytoplasmic expression and scattered nuclear expression. Figure 2 Beclin-1 expression patterns in malignant CMTs. (A) High cytoplasmic and high nuclear expression pattern. (B) High cytoplasmic and low nuclear expression pattern. (C) Low cytoplasmic and high nuclear expression pattern. (D) Low cytoplasmic and low nuclear expression pattern. Figure 3 Comparison of Q scores of Beclin-1 in normal mammary glands and malignant CMTs. A. Cytoplasmic expression in normal mammary glands. B. Cytoplasmic expression in malignant CMTs. C. Nuclear expression in normal mammary glands. D. Nuclear expression in malignant CMTs. The middle lines of boxes show the median value, whereas top and bottom lines of boxes represent interquartile range. The ends of whiskers represent 10th and 90th percentiles. The outliers are indicated by open circles. Association of Beclin-1 expression in cancer cells and clinicopathologic characteristics The associations between Beclin-1 expression patterns and clinicopathologic variables are shown in Table 1. The median value of the Q score of cytoplasmic Beclin-1 expression in malignant CMTs was 60. Using the median value as a cutoff point, 40 cases (57.14%) were classified as low cytoplasmic expression, whereas 30 cases (42.86%) were classified as high cytoplasmic expression. Low cytoplasmic expression (Q score 60) of Beclin-1 was associated with older age, lower degree of tubular formation, increased mitotic activity, higher histologic grade, and extensive necrosis. The median value of the nuclear Q score in malignant CMTs was 10. In total, 28 cases (40%) were sub-grouped into low nuclear expression, and 42 cases (60%) were sub-grouped into high nuclear expression. Low nuclear expression (Q score 10) of Beclin-1 was linked to older age, lower degree of tubular formation, extensive necrosis, and negative for Her2/neu overexpression. Beclin-1 cytoplasmic expression was linked significantly with nuclear expression (p = 0.003) (Table 2). Table 2 Association of nuclear expression and cytoplasmic expression of Beclin-1 in 70 cases of malignant CMTs Nuclear expression Low High p value Cytoplasmic expression Low 22 (31.43%) 18 (25.71%) 0.003 High 6 (8.57%) 24 (34.29%) * p < 0.05. Survival analysis The mean follow-up time was 21 ± 18.72 months. Univariate survival analysis using the Kaplan-Meir method revealed that age, tumor size, tubular formation, nuclear pleomorphism, mitotic count, histologic grade, lymphovascular invasion, necrosis, and Beclin-1 cytoplasmic expression were significant prognostic factors for overall survival (Table 3). Figure 4 shows the Kaplan-Meier curves of cumulative overall survival probability in relation to the Beclin-1 expression of cancer cells. Patients with low cytoplasmic expression showed poorer overall survival rate (p < 0.001). The difference of overall survival rate between high and low nuclear expressions was not statistically significant (p = 0.074). Multivariate survival analysis using the Cox proportional hazard regression method revealed that tumor size, tubular formation, and Beclin-1 cytoplasmic expression were independent prognostic factors for malignant CMTs (Table 4). Table 3 Univariate analysis of clinicopathologic variables for ovarall survival rate Variables p value Age (> 11 years vs. 11 years) 0.001* Tumor size (> 3 cm vs. 3 cm) 0.042* Tubular formation (> 10% vs. 10%) <0.001* Nuclear pleomorphism (high vs. low) 0.044* Mitotic count (> 11 vs. 10/10 HPFs) 0.005* Histologic grade (3 vs. 2 and 1) <0.001* Lymphovascular invasion (yes vs. no) <0.001* Necrosis (extensive vs. limited/no) 0.004* Estrogen receptor (negative vs. positive) 0.180 Her2/neu overexpression (positive vs. negative) 0.348 Beclin-1 cytoplasmic expression (low vs. high) <0.001* Beclin-1 nuclear expression (low vs. high) 0.074 * p < 0.05. Figure 4 Kaplan-Meier curves of overall survival rate in 70 cases of malignant CMTs. (A) Curves for patients with low and high cytoplasmic Beclin-1 expressions. (B) Curves for patients with low and high nuclear Beclin-1 expressions. Table 4 Multivariate Cox proportional hazard analysis of clinicopathologic variables for ovarall survival rate Multivariate analysis Hazard ratio 95% confidence interval p value Age (> 11 years vs. 11 years) 1.460 0.683 - 3.124 0.329 Tumor size (> 3 cm vs. 3 cm) 3.038 1.298 - 7.108 0.010* Tubular formation (> 10% vs. 10%) 0.364 0.167 - 0.794 0.011* Nuclear pleomorphism (high vs. low) 1.001 0.445 - 2.253 0.997 Mitotic count (> 11 vs. 10/10 HPFs) 0.881 0.166 - 4.676 0.882 Histologic grade (3 vs. 2 and 1) 0.627 0.088 - 4.486 0.642 Lymphovascular invasion (yes vs. no) 2.286 0.887 - 5.887 0.087 Necrosis (extensive vs. limited/no) 0.977 0.473 - 2.019 0.951 Estrogen receptor (negative vs. positive) 0.499 0.232 - 1.076 0.076 Her2/neu overexpression (positive vs. negative) 2.433 0.950 - 6.234 0.064 Beclin-1 cytoplasmic expression (low vs. high) 2.752 1.211 - 6.256 0.016* Beclin-1 nuclear expression (low vs. high) 1.471 0.667 - 3.243 0.339 * p < 0.05. Discussion Autophagy, an essential catabolic mechanism, is also involved in tumor initiation and progression. Recent studies have revealed that the expression of Beclin-1 is decreased in various human cancer types, such as breast [10], cervical [11], esophageal [12], lung cancers [13, 14], hepatocellular carcinoma [15], and cutaneous melanoma [16]. However, Beclin-1 expression was reported to be increased in human colon, gastric, and pancreatic cancers, in contrast to their normal counterparts [17, 18]. The mechanism of aberrance of Beclin-1 expression in different types of cancers is largely unknown. These variable results imply that autophagic activity is specific in different organs and histologic types. They also indicate that autophagy may either induce or inhibit tumor cell survival. In this study, we compared the Beclin-1 expression in malignant CMTs and surrounding normal mammary glands. Cytoplasmic expression of cancer cells was significantly lower than that of normal mammary glands. Decreased expression of Beclin-1 was associated with some aggressive histologic features. These findings were similar to those of human breast cancer [7, 10]. Malignant CMT has similar epidemiologic, histologic, clinical, and prognostic features to human breast cancer. Our results imply that the autophagic activities in canine and human mammary glands may also be coincidental. Further comparative studies of autophagy may be beneficial to both human beings and dogs. The subcellular localization of Beclin-1 was mainly reported at the cytoplasm and/or membrane, and the nuclear expression pattern was also documented [19, 20]. The leucine-rich nuclear export signal of Beclin-1 is essential for autophagic growth control and tumor suppression [21]. Our study disclosed that nuclear expression is associated with cytoplasmic expression. Lower nuclear expression is also related to unfavorable clinicopathologic features. The relationship between the expression pattern of Beclin-1 and the prognosis was controversial in studies of human cancer. Loss of Beclin-1 was linked to poorer survival rate in stage III colon cancer [19], esophageal squamous cell carcinoma [12], hepatocellular carcinoma [15], intrahepatic cholangiocarcinoma [22], pancreatic ductal adenocarcinoma [18], chondrosarcoma [23], and several types of lymphoma [2426]. High Beclin-1 expression was connected to poor prognosis in endometrial adenocarcinoma [27] and nasopharyngeal carcinoma in humans [28]. Koukourakis et al. found that extensive overexpression and underexpression of Beclin-1 was associated with poor overall survival in human patients with colon cancer [20]. They noted that the nuclear expression of Beclin-1 was not related to the prognosis. These results indicate that Beclin-1 may either induce or inhibit tumor cell survival. Our results support the hypothesis that Beclin-1 functions as a tumor suppressor protein in malignant CMTs. The mechanisms by which autophagy suppresses and promotes carcinogenesis are not yet completely understood. Both autophagy promoters and autophagy inhibitors are clinically effective in cancer treatment [2932]. The autophagic tumor stroma model of cancer proposed by Martinez-Outschoorn et al. attempted to resolve the paradox [33]. In this model, cancer cells use oxidative stress to induce autophagy in the tumor environment, whereas the autophagic tumor stromal cells produce recycled nutrients to promote the growth of cancer cells [34]. Sanchez et al. discovered that the mesenchymal stem cell-derived stromal cells in human breast cancer showed upregulation of Beclin-1 and other autophagic markers [35]. However, this model may not explain the upregulation of autophagy-related proteins in some human cancer cells. Our study and other previous researches did not find a specific immunohistochemical staining pattern of Beclin-1 in cancer-associated stromal cells. Moreover, the Beclin-1 independent autophagic process may also be considered. More proteomic-based studies should be performed to clarify the functions of autophagy-related proteins in cancer and cancer-associated stromal cells. Conclusions We analyzed the Beclin-1 expression pattern in normal mammary glands and malignant CMTs. We found that the loss of Beclin-1 expression is associated with aggressive clinicohistologic features and poor overall survival. Our results suggest that Beclin-1 plays a significant role in tumor progression and can be a potential therapeutic target for malignant CMTs in the future. Methods Patients and tissue samples Formalin-fixed, paraffin-embedded tissue samples from 70 female dogs with primary malignant CMTs were analyzed in this study. The 70 dogs included 21 mongrels, 19 Maltese, 7 Shih-Tzus, 6 poodles, 4 Cocker spaniels, 4 Schnauzers, 4 Yorkshire terriers, 2 Labrador retrievers, 1 French spaniel, 1 Pomeranian, and 1 spitz. All of these specimens were surgically resected at National Taiwan University Veterinary Hospital from 2005 to 2011. Patients who received chemotherapy before or after surgery were excluded from this study. All cases were pathologically diagnosed with primary malignant CMTs at the School of Veterinary Medicine, National Taiwan University. Information such as age, breed, status of ovario-hysterectomy, and tumor size of patients was obtained from medical records. Follow-up data were obtained from medical records and by telephone contact with the dog owners. Overall survival was defined as the time between surgery and death. Pathologic examination Routine hematoxylin and eosin (HE) staining was performed for histologic assessment. The histologic type was assessed according to the diagnostic criteria of the World Health Organization Histological Classification of Mammary Tumors of the Dog and Cat [36]. The tumors were graded based on Nottingham Modification of the Bloom-Richardson system on HE-stained sections [37, 38]. The grading system combined 3 histopathologic features: tubular formation, nuclear pleomorphism, and mitotic counts. Each feature was scored 1 to 3 points. The histologic grade was according to the final scores as follows: 3, 4, or 5 points as grade I (well differentiated); 6 or 7 points as grade II (moderately differentiated); and 8 or 9 points as grade III (poorly differentiated). Necrotic areas of more than 10% of the tissue section were regarded as “extensive,” whereas the remaining cases were recorded as “limited/no” necrosis. The presence of cancer cells within vascular channels of the primary tumor was detected in HE–stained sections. Immunohistochemical staining Tissue blocks containing representative cancer areas with surrounding normal breast tissue were selected from every case. Immunohistochemical staining was performed on 4-μm-thick tissue sections using the Leica Bond-Max autostainer (Leica microsystem) according to the manufacturer’s instructions with minor modifications. Sections were deparaffinized by the Bond Dewax Solution (Leica Microsystems). Heat-induced antigen retrieval was achieved using the Bond Epitope Retrieval Solution 2 (Leica Microsystems) for 20 min at 100°C. Endogenous peroxidase activity was blocked by incubation of tissue with the Novocastra Peroxidase Block for 5 min. The slides were then incubated for 30 min at room temperature with a primary rabbit polyclonal antibody against Beclin-1 (1: 400, Abcam, UK), estrogen receptor (1:50, clone 6 F11, Novocastra, UK), and Her2 (1:50, clone CB11, Novocastra, UK). Diaminobenzidine-tetrahydrochloride (DAB) was used as the substrate to detect antigen-antibody binding. Sections were counterstained with hematoxylin. Immunohistochemical evaluation For Beclin-1, the intensity, percentage, and sub-localization of immunohistochemical stains in cancer cells and cancer-adjacent normal mammary glands of each case were recorded. The normal canine mammary ductal epithelium from dogs without mammary neoplasia was used as positive control, and staining with omission of primary antibody was performed as negative control. The intensity and percentage of positively stained cells were counted at 10 high-powered fields (400 ×). The intensity of staining was recorded as 0, 1, 2, and 3 standing for negative, weak, moderate and strong staining, respectively. The percentage of positive cells was scored from 0% to 100%. The results of cytoplasmic and nuclear Beclin-1 expressions were scored by quick score (Q), which is obtained by multiplying the percentage of positive cells (P) by the intensity (I) (Q = P × I; maximum = 300) [39]. The median value of the Q score in cancer cells was used as a cutoff point, and the cases were sub-grouped into “low expression” and “high expression”. The Her2 expression was scored according to the American Society of Clinical Oncology/College of American Pathologists guidelines (0 = no staining or membrane staining in fewer than 10% of tumor cells; 1 + = faint, barely perceptible membrane staining in more than 10% of tumor cells; 2 + = weak to moderate complete membrane staining observed in more than 10% of tumor cells or strong complete membrane staining in less than 30% of tumor cells; 3 + = strong and complete membrane staining in more than 30% tumor cells) [40]. In this study, Her2 positive was defined as a score of 3+, whereas the rest were regarded as negative. For estrogen receptor, nuclear staining more than 10% of cancer cells were classified as positive, while the others were classified as negative. Immunohistochemical staining was evaluated by two pathologists independently without knowledge of clinical outcomes of the patients. Conflicting results were resolved at multi-headed microscope. Statistical analysis The Wilcoxon signed-rank test was used to analyze Beclin-1 expression in paired normal mammary glands and cancer cells. The chi-square test was used to evaluate the association of Beclin-1 expression with clinicopathologic features of malignant CMTs. Curves for overall survival were drawn using the Kaplan-Meier method, and the differences of survival rate were compared using the log-rank test for univariate survival analysis. The Cox proportion hazard regression model was used for multivariate survival analysis of prognostic factors. A p value of less than 0.05 was considered statistically significant. 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3.1 使用Java运算符运算符以一个或多个自变量为基础，可生成一个新值。自变量采用与原始方法调用不同的一种形式，但效果是相同的。根据以前写程序的经验，运算符的常规概念应该不难理解。加号（+）、减号和负号（-）、乘号（）、除号（/）以及等号（=）的用法与其他所有编程语言都是类似的。所有运算符都能根据自己的运算对象生成一个值。除此以外，一个运算符可改变运算对象的值，这叫作“副作用”（Side Effect）。运算符最常见的用途就是修改自己的运算对象，从而产生副作用。但要注意生成的值亦可由没有副作用的运算符生成。几乎所有运算符都只能操作“主类型”（Primitives）。唯一的例外是“=”、“==”和“!=”，它们能操作所有对象（也是对象易令人混淆的一个地方）。除此以外，String类支持“+”和“+=”。 3.1.1 优先级运算符的优先级决定了存在多个运算符时一个表达式各部分的计算顺序。Java对计算顺序作出了特别的规定。其中，最简单的规则就是乘法和除法在加法和减法之前完成。程序员经常都会忘记其他优先级规则，所以应该用括号明确规定计算顺序。例如： A = X + Y - 2/2 + Z; 为上述表达式加上括号后，就有了一个不同的含义。 A = X + (Y - 2)/(2 + Z); 3.1.2 赋值赋值是用等号运算符（=）进行的。它的意思是“取得右边的值，把它复制到左边”。右边的值可以是任何常数、变量或者表达式，只要能产生一个值就行。但左边的值必须是一个明确的、已命名的变量。也就是说，它必须有一个物理性的空间来保存右边的值。举个例子来说，可将一个常数赋给一个变量（A=4;），但不可将任何东西赋给一个常数（比如不能4=A）。对主数据类型的赋值是非常直接的。由于主类型容纳了实际的值，而且并非指向一个对象的句柄，所以在为其赋值的时候，可将来自一个地方的内容复制到另一个地方。例如，假设为主类型使用“A=B”，那么B处的内容就复制到A。若接着又修改了A，那么B根本不会受这种修改的影响。作为一名程序员，这应成为自己的常识。但在为对象“赋值”的时候，情况却发生了变化。对一个对象进行操作时，我们真正操作的是它的句柄。所以倘若“从一个对象到另一个对象”赋值，实际就是将句柄从一个地方复制到另一个地方。这意味着假若为对象使用“C=D”，那么C和D最终都会指向最初只有D才指向的那个对象。下面这个例子将向大家阐示这一点。这里有一些题外话。在后面，大家在代码示例里看到的第一个语句将是“package 03”使用的“package”语句，它代表本书第3章。本书每一章的第一个代码清单都会包含象这样的一个“package”（封装、打包、包裹）语句，它的作用是为那一章剩余的代码建立章节编号。在第17章，大家会看到第3章的所有代码清单（除那些有不同封装名称的以外）都会自动置入一个名为c03的子目录里；第4章的代码置入c04；以此类推。所有这些都是通过第17章展示的CodePackage.java程序实现的；“封装”的基本概念会在第5章进行详尽的解释。就目前来说，大家只需记住象“package 03”这样的形式只是用于为某一章的代码清单建立相应的子目录。为运行程序，必须保证在classpath里包含了我们安装本书源码文件的根目录（那个目录里包含了c02，c03c，c04等等子目录）。对于Java后续的版本（1.1.4和更高版本），如果您的main()用package语句封装到一个文件里，那么必须在程序名前面指定完整的包裹名称，否则不能运行程序。在这种情况下，命令行是： java c03.Assignment 运行位于一个“包裹”里的程序时，随时都要注意这方面的问题。下面是例子： //: Assignment.java // Assignment with objects is a bit tricky package c03; class Number { int i; } public class Assignment { public static void main(String[] args) { Number n1 = new Number(); Number n2 = new Number(); n1.i = 9; n2.i = 47; System.out.println("1: n1.i: " + n1.i + ", n2.i: " + n2.i); n1 = n2; System.out.println("2: n1.i: " + n1.i + ", n2.i: " + n2.i); n1.i = 27; System.out.println("3: n1.i: " + n1.i + ", n2.i: " + n2.i); } } ///:~ Number类非常简单，它的两个实例（n1和n2）是在main()里创建的。每个Number中的i值都赋予了一个不同的值。随后，将n2赋给n1，而且n1发生改变。在许多程序设计语言中，我们都希望n1和n2任何时候都相互独立。但由于我们已赋予了一个句柄，所以下面才是真实的输出： 1: n1.i: 9, n2.i: 47 2: n1.i: 47, n2.i: 47 3: n1.i: 27, n2.i: 27 看来改变n1的同时也改变了n2！这是由于无论n1还是n2都包含了相同的句柄，它指向相同的对象（最初的句柄位于n1内部，指向容纳了值9的一个对象。在赋值过程中，那个句柄实际已经丢失；它的对象会由“垃圾收集器”自动清除）。这种特殊的现象通常也叫作“别名”，是Java操作对象的一种基本方式。但假若不愿意在这种情况下出现别名，又该怎么操作呢？可放弃赋值，并写入下述代码： n1.i = n2.i; 这样便可保留两个独立的对象，而不是将n1和n2绑定到相同的对象。但您很快就会意识到，这样做会使对象内部的字段处理发生混乱，并与标准的面向对象设计准则相悖。由于这并非一个简单的话题，所以留待第12章详细论述，那一章是专门讨论别名的。其时，大家也会注意到对象的赋值会产生一些令人震惊的效果。 1. 方法调用中的别名处理将一个对象传递到方法内部时，也会产生别名现象。 //: PassObject.java // Passing objects to methods can be a bit tricky class Letter { char c; } public class PassObject { static void f(Letter y) { y.c = 'z'; } public static void main(String[] args) { Letter x = new Letter(); x.c = 'a'; System.out.println("1: x.c: " + x.c); f(x); System.out.println("2: x.c: " + x.c); } } ///:~ 在许多程序设计语言中，f()方法表面上似乎要在方法的作用域内制作自己的自变量Letter y的一个副本。但同样地，实际传递的是一个句柄。所以下面这个程序行： y.c = 'z'; ` 实际改变的是f()之外的对象。输出结果如下： 1: x.c: a 2: x.c: z 别名和它的对策是非常复杂的一个问题。尽管必须等至第12章才可获得所有答案，但从现在开始就应加以重视，以便提早发现它的缺点。 3.1.3 算术运算符 Java的基本算术运算符与其他大多数程序设计语言是相同的。其中包括加号（+）、减号（-）、除号（/）、乘号（）以及模数（%，从整数除法中获得余数）。整数除法会直接砍掉小数，而不是进位。 Java也用一种简写形式进行运算，并同时进行赋值操作。这是由等号前的一个运算符标记的，而且对于语言中的所有运算符都是固定的。例如，为了将4加到变量x，并将结果赋给x，可用：x+=4。下面这个例子展示了算术运算符的各种用法： //: MathOps.java // Demonstrates the mathematical operators import java.util.; public class MathOps { // Create a shorthand to save typing: static void prt(String s) { System.out.println(s); } // shorthand to print a string and an int: static void pInt(String s, int i) { prt(s + " = " + i); } // shorthand to print a string and a float: static void pFlt(String s, float f) { prt(s + " = " + f); } public static void main(String[] args) { // Create a random number generator, // seeds with current time by default: Random rand = new Random(); int i, j, k; // '%' limits maximum value to 99: j = rand.nextInt() % 100; k = rand.nextInt() % 100; pInt("j",j); pInt("k",k); i = j + k; pInt("j + k", i); i = j - k; pInt("j - k", i); i = k / j; pInt("k / j", i); i = k j; pInt("k * j", i); i = k % j; pInt("k % j", i); j %= k; pInt("j %= k", j); // Floating-point number tests: float u,v,w; // applies to doubles, too v = rand.nextFloat(); w = rand.nextFloat(); pFlt("v", v); pFlt("w", w); u = v + w; pFlt("v + w", u); u = v - w; pFlt("v - w", u); u = v * w; pFlt("v * w", u); u = v / w; pFlt("v / w", u); // the following also works for // char, byte, short, int, long, // and double: u += v; pFlt("u += v", u); u -= v; pFlt("u -= v", u); u = v; pFlt("u = v", u); u /= v; pFlt("u /= v", u); } } ///:~ 我们注意到的第一件事情就是用于打印（显示）的一些快捷方法：prt()方法打印一个String；pInt()先打印一个String，再打印一个int；而pFlt()先打印一个String，再打印一个float。当然，它们最终都要用System.out.println()结尾。为生成数字，程序首先会创建一个Random（随机）对象。由于自变量是在创建过程中传递的，所以Java将当前时间作为一个“种子值”，由随机数生成器利用。通过Random对象，程序可生成许多不同类型的随机数字。做法很简单，只需调用不同的方法即可：nextInt()，nextLong()，nextFloat()或者nextDouble()。若随同随机数生成器的结果使用，模数运算符（%）可将结果限制到运算对象减1的上限（本例是99）之下。 1. 一元加、减运算符一元减号（-）和一元加号（+）与二元加号和减号都是相同的运算符。根据表达式的书写形式，编译器会自动判断使用哪一种。例如下述语句： x = -a; 它的含义是显然的。编译器能正确识别下述语句： x = a * -b; 但读者会被搞糊涂，所以最好更明确地写成： x = a * (-b); 一元减号得到的运算对象的负值。一元加号的含义与一元减号相反，虽然它实际并不做任何事情。 3.1.4 自动递增和递减和C类似，Java提供了丰富的快捷运算方式。这些快捷运算可使代码更清爽，更易录入，也更易读者辨读。两种很不错的快捷运算方式是递增和递减运算符（常称作“自动递增”和“自动递减”运算符）。其中，递减运算符是“--”，意为“减少一个单位”；递增运算符是“++”，意为“增加一个单位”。举个例子来说，假设A是一个int（整数）值，则表达式++A就等价于（A = A + 1）。递增和递减运算符结果生成的是变量的值。对每种类型的运算符，都有两个版本可供选用；通常将其称为“前缀版”和“后缀版”。“前递增”表示++运算符位于变量或表达式的前面；而“后递增”表示++运算符位于变量或表达式的后面。类似地，“前递减”意味着--运算符位于变量或表达式的前面；而“后递减”意味着--运算符位于变量或表达式的后面。对于前递增和前递减（如++A或--A），会先执行运算，再生成值。而对于后递增和后递减（如A++或A--），会先生成值，再执行运算。下面是一个例子： //: AutoInc.java // Demonstrates the ++ and -- operators public class AutoInc { public static void main(String[] args) { int i = 1; prt("i : " + i); prt("++i : " + ++i); // Pre-increment prt("i++ : " + i++); // Post-increment prt("i : " + i); prt("--i : " + --i); // Pre-decrement prt("i-- : " + i--); // Post-decrement prt("i : " + i); } static void prt(String s) { System.out.println(s); } } ///:~ 该程序的输出如下： i : 1 ++i : 2 i++ : 2 i : 3 --i : 2 i-- : 2 i : 1 从中可以看到，对于前缀形式，我们在执行完运算后才得到值。但对于后缀形式，则是在运算执行之前就得到值。它们是唯一具有“副作用”的运算符（除那些涉及赋值的以外）。也就是说，它们会改变运算对象，而不仅仅是使用自己的值。递增运算符正是对“C++”这个名字的一种解释，暗示着“超载C的一步”。在早期的一次Java演讲中，Bill Joy（始创人之一）声称“Java=C++--”（C加加减减），意味着Java已去除了C++一些没来由折磨人的地方，形成一种更精简的语言。正如大家会在这本书中学到的那样，Java的许多地方都得到了简化，所以Java的学习比C++更容易。 3.1.5 关系运算符关系运算符生成的是一个“布尔”（Boolean）结果。它们评价的是运算对象值之间的关系。若关系是真实的，关系表达式会生成true（真）；若关系不真实，则生成false（假）。关系运算符包括小于（<）、大于（>）、小于或等于（<=）、大于或等于（>=）、等于（==）以及不等于（!=）。等于和不等于适用于所有内建的数据类型，但其他比较不适用于boolean类型。 1. 检查对象是否相等关系运算符==和!=也适用于所有对象，但它们的含义通常会使初涉Java领域的人找不到北。下面是一个例子： //: Equivalence.java public class Equivalence { public static void main(String[] args) { Integer n1 = new Integer(47); Integer n2 = new Integer(47); System.out.println(n1 == n2); System.out.println(n1 != n2); } } ///:~ 其中，表达式System.out.println(n1 == n2)可打印出内部的布尔比较结果。一般人都会认为输出结果肯定先是true，再是false，因为两个Integer对象都是相同的。但尽管对象的内容相同，句柄却是不同的，而==和!=比较的正好就是对象句柄。所以输出结果实际上先是false，再是true。这自然会使第一次接触的人感到惊奇。若想对比两个对象的实际内容是否相同，又该如何操作呢？此时，必须使用所有对象都适用的特殊方法equals()。但这个方法不适用于“主类型”，那些类型直接使用==和!=即可。下面举例说明如何使用： //: EqualsMethod.java public class EqualsMethod { public static void main(String[] args) { Integer n1 = new Integer(47); Integer n2 = new Integer(47); System.out.println(n1.equals(n2)); } } ///:~ 正如我们预计的那样，此时得到的结果是true。但事情并未到此结束！假设您创建了自己的类，就象下面这样： //: EqualsMethod2.java class Value { int i; } public class EqualsMethod2 { public static void main(String[] args) { Value v1 = new Value(); Value v2 = new Value(); v1.i = v2.i = 100; System.out.println(v1.equals(v2)); } } ///:~ 此时的结果又变回了false！这是由于equals()的默认行为是比较句柄。所以除非在自己的新类中改变了equals()，否则不可能表现出我们希望的行为。不幸的是，要到第7章才会学习如何改变行为。但要注意equals()的这种行为方式同时或许能够避免一些“灾难”性的事件。大多数Java类库都实现了equals()，所以它实际比较的是对象的内容，而非它们的句柄。 3.1.6 逻辑运算符逻辑运算符AND（&&）、OR（\|\|）以及NOT（!）能生成一个布尔值（true或false）——以自变量的逻辑关系为基础。下面这个例子向大家展示了如何使用关系和逻辑运算符。 //: Bool.java // Relational and logical operators import java.util.; public class Bool { public static void main(String[] args) { Random rand = new Random(); int i = rand.nextInt() % 100; int j = rand.nextInt() % 100; prt("i = " + i); prt("j = " + j); prt("i > j is " + (i > j)); prt("i < j is " + (i < j)); prt("i >= j is " + (i >= j)); prt("i <= j is " + (i <= j)); prt("i == j is " + (i == j)); prt("i != j is " + (i != j)); // Treating an int as a boolean is // not legal Java //! prt("i && j is " + (i && j)); //! prt("i \|\| j is " + (i \|\| j)); //! prt("!i is " + !i); prt("(i < 10) && (j < 10) is " + ((i < 10) && (j < 10)) ); prt("(i < 10) \|\| (j < 10) is " + ((i < 10) \|\| (j < 10)) ); } static void prt(String s) { System.out.println(s); } } ///:~ 只可将AND，OR或NOT应用于布尔值。与在C及C++中不同，不可将一个非布尔值当作布尔值在逻辑表达式中使用。若这样做，就会发现尝试失败，并用一个“//!”标出。然而，后续的表达式利用关系比较生成布尔值，然后对结果进行逻辑运算。输出列表看起来象下面这个样子： i = 85 j = 4 i > j is true i < j is false i >= j is true i <= j is false i == j is false i != j is true (i < 10) && (j < 10) is false (i < 10) \|\| (j < 10) is true 注意若在预计为String值的地方使用，布尔值会自动转换成适当的文本形式。在上述程序中，可将对int的定义替换成除boolean以外的其他任何主数据类型。但要注意，对浮点数字的比较是非常严格的。即使一个数字仅在小数部分与另一个数字存在极微小的差异，仍然认为它们是“不相等”的。即使一个数字只比零大一点点（例如2不停地开平方根），它仍然属于“非零”值。 1. 短路操作逻辑运算符时，我们会遇到一种名为“短路”的情况。这意味着只有明确得出整个表达式真或假的结论，才会对表达式进行逻辑求值。因此，一个逻辑表达式的所有部分都有可能不进行求值： //: ShortCircuit.java // Demonstrates short-circuiting behavior // with logical operators. public class ShortCircuit { static boolean test1(int val) { System.out.println("test1(" + val + ")"); System.out.println("result: " + (val < 1)); return val < 1; } static boolean test2(int val) { System.out.println("test2(" + val + ")"); System.out.println("result: " + (val < 2)); return val < 2; } static boolean test3(int val) { System.out.println("test3(" + val + ")"); System.out.println("result: " + (val < 3)); return val < 3; } public static void main(String[] args) { if(test1(0) && test2(2) && test3(2)) System.out.println("expression is true"); else System.out.println("expression is false"); } } ///:~ 每次测试都会比较自变量，并返回真或假。它不会显示与准备调用什么有关的资料。测试在下面这个表达式中进行： if(test1(0)) && test2(2) && test3(2)) 很自然地，你也许认为所有这三个测试都会得以执行。但希望输出结果不至于使你大吃一惊： if(test1(0) && test2(2) && test3(2)) 第一个测试生成一个true结果，所以表达式求值会继续下去。然而，第二个测试产生了一个false结果。由于这意味着整个表达式肯定为false，所以为什么还要继续剩余的表达式呢？这样做只会徒劳无益。事实上，“短路”一词的由来正种因于此。如果一个逻辑表达式的所有部分都不必执行下去，那么潜在的性能提升将是相当可观的。 3.1.7 按位运算符按位运算符允许我们操作一个整数主数据类型中的单个“比特”，即二进制位。按位运算符会对两个自变量中对应的位执行布尔代数，并最终生成一个结果。按位运算来源于C语言的低级操作。我们经常都要直接操纵硬件，需要频繁设置硬件寄存器内的二进制位。Java的设计初衷是嵌入电视顶置盒内，所以这种低级操作仍被保留下来了。然而，由于操作系统的进步，现在也许不必过于频繁地进行按位运算。若两个输入位都是1，则按位AND运算符（&）在输出位里生成一个1；否则生成0。若两个输入位里至少有一个是1，则按位OR运算符（\|）在输出位里生成一个1；只有在两个输入位都是0的情况下，它才会生成一个0。若两个输入位的某一个是1，但不全都是1，那么按位XOR（^，异或）在输出位里生成一个1。按位NOT（~，也叫作“非”运算符）属于一元运算符；它只对一个自变量进行操作（其他所有运算符都是二元运算符）。按位NOT生成与输入位的相反的值——若输入0，则输出1；输入1，则输出0。按位运算符和逻辑运算符都使用了同样的字符，只是数量不同。因此，我们能方便地记忆各自的含义：由于“位”是非常“小”的，所以按位运算符仅使用了一个字符。按位运算符可与等号（=）联合使用，以便合并运算及赋值：&=，\|=和^=都是合法的（由于~是一元运算符，所以不可与=联合使用）。我们将boolean（布尔）类型当作一种“单位”或“单比特”值对待，所以它多少有些独特的地方。我们可执行按位AND，OR和XOR，但不能执行按位NOT（大概是为了避免与逻辑NOT混淆）。对于布尔值，按位运算符具有与逻辑运算符相同的效果，只是它们不会中途“短路”。此外，针对布尔值进行的按位运算为我们新增了一个XOR逻辑运算符，它并未包括在“逻辑”运算符的列表中。在移位表达式中，我们被禁止使用布尔运算，原因将在下面解释。 3.1.8 移位运算符移位运算符面向的运算对象也是二进制的“位”。可单独用它们处理整数类型（主类型的一种）。左移位运算符（<<）能将运算符左边的运算对象向左移动运算符右侧指定的位数（在低位补0）。“有符号”右移位运算符（>>）则将运算符左边的运算对象向右移动运算符右侧指定的位数。“有符号”右移位运算符使用了“符号扩展”：若值为正，则在高位插入0；若值为负，则在高位插入1。Java也添加了一种“无符号”右移位运算符（>>>），它使用了“零扩展”：无论正负，都在高位插入0。这一运算符是C或C++没有的。若对char，byte或者short进行移位处理，那么在移位进行之前，它们会自动转换成一个int。只有右侧的5个低位才会用到。这样可防止我们在一个int数里移动不切实际的位数。若对一个long值进行处理，最后得到的结果也是long。此时只会用到右侧的6个低位，防止移动超过long值里现成的位数。但在进行“无符号”右移位时，也可能遇到一个问题。若对byte或short值进行右移位运算，得到的可能不是正确的结果（Java 1.0和Java 1.1特别突出）。它们会自动转换成int类型，并进行右移位。但“零扩展”不会发生，所以在那些情况下会得到-1的结果。可用下面这个例子检测自己的实现方案： //: URShift.java // Test of unsigned right shift public class URShift { public static void main(String[] args) { int i = -1; i >>>= 10; System.out.println(i); long l = -1; l >>>= 10; System.out.println(l); short s = -1; s >>>= 10; System.out.println(s); byte b = -1; b >>>= 10; System.out.println(b); } } ///:~ 移位可与等号（<<=或>>=或>>>=）组合使用。此时，运算符左边的值会移动由右边的值指定的位数，再将得到的结果赋回左边的值。下面这个例子向大家阐示了如何应用涉及“按位”操作的所有运算符，以及它们的效果： //: BitManipulation.java // Using the bitwise operators import java.util.; public class BitManipulation { public static void main(String[] args) { Random rand = new Random(); int i = rand.nextInt(); int j = rand.nextInt(); pBinInt("-1", -1); pBinInt("+1", +1); int maxpos = 2147483647; pBinInt("maxpos", maxpos); int maxneg = -2147483648; pBinInt("maxneg", maxneg); pBinInt("i", i); pBinInt("~i", ~i); pBinInt("-i", -i); pBinInt("j", j); pBinInt("i & j", i & j); pBinInt("i \| j", i \| j); pBinInt("i ^ j", i ^ j); pBinInt("i << 5", i << 5); pBinInt("i >> 5", i >> 5); pBinInt("(~i) >> 5", (~i) >> 5); pBinInt("i >>> 5", i >>> 5); pBinInt("(~i) >>> 5", (~i) >>> 5); long l = rand.nextLong(); long m = rand.nextLong(); pBinLong("-1L", -1L); pBinLong("+1L", +1L); long ll = 9223372036854775807L; pBinLong("maxpos", ll); long lln = -9223372036854775808L; pBinLong("maxneg", lln); pBinLong("l", l); pBinLong("~l", ~l); pBinLong("-l", -l); pBinLong("m", m); pBinLong("l & m", l & m); pBinLong("l \| m", l \| m); pBinLong("l ^ m", l ^ m); pBinLong("l << 5", l << 5); pBinLong("l >> 5", l >> 5); pBinLong("(~l) >> 5", (~l) >> 5); pBinLong("l >>> 5", l >>> 5); pBinLong("(~l) >>> 5", (~l) >>> 5); } static void pBinInt(String s, int i) { System.out.println( s + ", int: " + i + ", binary: "); System.out.print(" "); for(int j = 31; j >=0; j--) if(((1 << j) & i) != 0) System.out.print("1"); else System.out.print("0"); System.out.println(); } static void pBinLong(String s, long l) { System.out.println( s + ", long: " + l + ", binary: "); System.out.print(" "); for(int i = 63; i >=0; i--) if(((1L << i) & l) != 0) System.out.print("1"); else System.out.print("0"); System.out.println(); } } ///:~ 程序末尾调用了两个方法：pBinInt()和pBinLong()。它们分别操作一个int和long值，并用一种二进制格式输出，同时附有简要的说明文字。目前，可暂时忽略它们具体的实现方案。大家要注意的是System.out.print()的使用，而不是System.out.println()。print()方法不会产生一个新行，以便在同一行里罗列多种信息。除展示所有按位运算符针对int和long的效果之外，本例也展示了int和long的最小值、最大值、+1和-1值，使大家能体会它们的情况。注意高位代表正负号：0为正，1为负。下面列出int部分的输出： -1, int: -1, binary: 11111111111111111111111111111111 +1, int: 1, binary: 00000000000000000000000000000001 maxpos, int: 2147483647, binary: 01111111111111111111111111111111 maxneg, int: -2147483648, binary: 10000000000000000000000000000000 i, int: 59081716, binary: 00000011100001011000001111110100 ~i, int: -59081717, binary: 11111100011110100111110000001011 -i, int: -59081716, binary: 11111100011110100111110000001100 j, int: 198850956, binary: 00001011110110100011100110001100 i & j, int: 58720644, binary: 00000011100000000000000110000100 i \| j, int: 199212028, binary: 00001011110111111011101111111100 i ^ j, int: 140491384, binary: 00001000010111111011101001111000 i << 5, int: 1890614912, binary: 01110000101100000111111010000000 i >> 5, int: 1846303, binary: 00000000000111000010110000011111 (~i) >> 5, int: -1846304, binary: 11111111111000111101001111100000 i >>> 5, int: 1846303, binary: 00000000000111000010110000011111 (~i) >>> 5, int: 132371424, binary: 00000111111000111101001111100000 数字的二进制形式表现为“有符号2的补值”。 3.1.9 三元if-else运算符这种运算符比较罕见，因为它有三个运算对象。但它确实属于运算符的一种，因为它最终也会生成一个值。这与本章后一节要讲述的普通if-else语句是不同的。表达式采取下述形式：布尔表达式 ? 值0:值1 若“布尔表达式”的结果为true，就计算“值0”，而且它的结果成为最终由运算符产生的值。但若“布尔表达式”的结果为false，计算的就是“值1”，而且它的结果成为最终由运算符产生的值。当然，也可以换用普通的if-else语句（在后面介绍），但三元运算符更加简洁。尽管C引以为傲的就是它是一种简练的语言，而且三元运算符的引入多半就是为了体现这种高效率的编程，但假若您打算频繁用它，还是要先多作一些思量——它很容易就会产生可读性极差的代码。可将条件运算符用于自己的“副作用”，或用于它生成的值。但通常都应将其用于值，因为那样做可将运算符与if-else明确区别开。下面便是一个例子： static int ternary(int i) { return i < 10 ? i * 100 : i * 10; } 可以看出，假设用普通的if-else结构写上述代码，代码量会比上面多出许多。如下所示： static int alternative(int i) { if (i < 10) return i * 100; return i * 10; } 但第二种形式更易理解，而且不要求更多的录入。所以在挑选三元运算符时，请务必权衡一下利弊。 3.1.10 逗号运算符在C和C++里，逗号不仅作为函数自变量列表的分隔符使用，也作为进行后续计算的一个运算符使用。在Java里需要用到逗号的唯一场所就是for循环，本章稍后会对此详加解释。 3.1.11 字串运算符+ 这个运算符在Java里有一项特殊用途：连接不同的字串。这一点已在前面的例子中展示过了。尽管与+的传统意义不符，但用+来做这件事情仍然是非常自然的。在C++里，这一功能看起来非常不错，所以引入了一项“运算符过载”机制，以便C++程序员为几乎所有运算符增加特殊的含义。但非常不幸，与C++的另外一些限制结合，运算符过载成为一种非常复杂的特性，程序员在设计自己的类时必须对此有周到的考虑。与C++相比，尽管运算符过载在Java里更易实现，但迄今为止仍然认为这一特性过于复杂。所以Java程序员不能象C++程序员那样设计自己的过载运算符。我们注意到运用“String +”时一些有趣的现象。若表达式以一个String起头，那么后续所有运算对象都必须是字串。如下所示： int x = 0, y = 1, z = 2; String sString = "x, y, z "; System.out.println(sString + x + y + z); 在这里，Java编译程序会将x，y和z转换成它们的字串形式，而不是先把它们加到一起。然而，如果使用下述语句： System.out.println(x + sString); 那么早期版本的Java就会提示出错（以后的版本能将x转换成一个字串）。因此，如果想通过“加号”连接字串（使用Java的早期版本），请务必保证第一个元素是字串（或加上引号的一系列字符，编译能将其识别成一个字串）。 3.1.12 运算符常规操作规则使用运算符的一个缺点是括号的运用经常容易搞错。即使对一个表达式如何计算有丝毫不确定的因素，都容易混淆括号的用法。这个问题在Java里仍然存在。在C和C++中，一个特别常见的错误如下： while(x = y) { //... } 程序的意图是测试是否“相等”（==），而不是进行赋值操作。在C和C++中，若y是一个非零值，那么这种赋值的结果肯定是true。这样使可能得到一个无限循环。在Java里，这个表达式的结果并不是布尔值，而编译器期望的是一个布尔值，而且不会从一个int数值中转换得来。所以在编译时，系统就会提示出现错误，有效地阻止我们进一步运行程序。所以这个缺点在Java里永远不会造成更严重的后果。唯一不会得到编译错误的时候是x和y都为布尔值。在这种情况下，x = y属于合法表达式。而在上述情况下，则可能是一个错误。在C和C++里，类似的一个问题是使用按位AND和OR，而不是逻辑AND和OR。按位AND和OR使用两个字符之一（&或\|），而逻辑AND和OR使用两个相同的字符（&&或\|\|）。就象“=”和“==”一样，键入一个字符当然要比键入两个简单。在Java里，编译器同样可防止这一点，因为它不允许我们强行使用一种并不属于的类型。 3.1.13 造型运算符 “造型”（Cast）的作用是“与一个模型匹配”。在适当的时候，Java会将一种数据类型自动转换成另一种。例如，假设我们为浮点变量分配一个整数值，计算机会将int自动转换成float。通过造型，我们可明确设置这种类型的转换，或者在一般没有可能进行的时候强迫它进行。为进行一次造型，要将括号中希望的数据类型（包括所有修改符）置于其他任何值的左侧。下面是一个例子： void casts() { int i = 200; long l = (long)i; long l2 = (long)200; } 正如您看到的那样，既可对一个数值进行造型处理，亦可对一个变量进行造型处理。但在这儿展示的两种情况下，造型均是多余的，因为编译器在必要的时候会自动进行int值到long值的转换。当然，仍然可以设置一个造型，提醒自己留意，也使程序更清楚。在其他情况下，造型只有在代码编译时才显出重要性。在C和C++中，造型有时会让人头痛。在Java里，造型则是一种比较安全的操作。但是，若进行一种名为“缩小转换”（Narrowing Conversion）的操作（也就是说，脚本是能容纳更多信息的数据类型，将其转换成容量较小的类型），此时就可能面临信息丢失的危险。此时，编译器会强迫我们进行造型，就好象说：“这可能是一件危险的事情——如果您想让我不顾一切地做，那么对不起，请明确造型。”而对于“放大转换”（Widening conversion），则不必进行明确造型，因为新类型肯定能容纳原来类型的信息，不会造成任何信息的丢失。 Java允许我们将任何主类型“造型”为其他任何一种主类型，但布尔值（bollean）要除外，后者根本不允许进行任何造型处理。“类”不允许进行造型。为了将一种类转换成另一种，必须采用特殊的方法（字串是一种特殊的情况，本书后面会讲到将对象造型到一个类型“家族”里；例如，“橡树”可造型为“树”；反之亦然。但对于其他外来类型，如“岩石”，则不能造型为“树”）。 1. 字面值最开始的时候，若在一个程序里插入“字面值”（Literal），编译器通常能准确知道要生成什么样的类型。但在有些时候，对于类型却是暧昧不清的。若发生这种情况，必须对编译器加以适当的“指导”。方法是用与字面值关联的字符形式加入一些额外的信息。下面这段代码向大家展示了这些字符。 //: Literals.java class Literals { char c = 0xffff; // max char hex value byte b = 0x7f; // max byte hex value short s = 0x7fff; // max short hex value int i1 = 0x2f; // Hexadecimal (lowercase) int i2 = 0X2F; // Hexadecimal (uppercase) int i3 = 0177; // Octal (leading zero) // Hex and Oct also work with long. long n1 = 200L; // long suffix long n2 = 200l; // long suffix long n3 = 200; //! long l6(200); // not allowed float f1 = 1; float f2 = 1F; // float suffix float f3 = 1f; // float suffix float f4 = 1e-45f; // 10 to the power float f5 = 1e+9f; // float suffix double d1 = 1d; // double suffix double d2 = 1D; // double suffix double d3 = 47e47d; // 10 to the power } ///:~ 十六进制（Base 16）——它适用于所有整数数据类型——用一个前置的0x或0X指示。并在后面跟随采用大写或小写形式的0-9以及a-f。若试图将一个变量初始化成超出自身能力的一个值（无论这个值的数值形式如何），编译器就会向我们报告一条出错消息。注意在上述代码中，最大的十六进制值只会在char，byte以及short身上出现。若超出这一限制，编译器会将值自动变成一个int，并告诉我们需要对这一次赋值进行“缩小造型”。这样一来，我们就可清楚获知自己已超载了边界。八进制（Base 8）是用数字中的一个前置0以及0-7的数位指示的。在C，C++或者Java中，对二进制数字没有相应的“字面”表示方法。字面值后的尾随字符标志着它的类型。若为大写或小写的L，代表long；大写或小写的F，代表float；大写或小写的D，则代表double。指数总是采用一种我们认为很不直观的记号方法：1.39e-47f。在科学与工程学领域，“e”代表自然对数的基数，约等于2.718（Java一种更精确的double值采用Math.E的形式）。它在象“1.39×e的-47次方”这样的指数表达式中使用，意味着“1.39×2.718的-47次方”。然而，自FORTRAN语言发明后，人们自然而然地觉得e代表“10多少次幂”。这种做法显得颇为古怪，因为FORTRAN最初面向的是科学与工程设计领域。理所当然，它的设计者应对这样的混淆概念持谨慎态度（注释①）。但不管怎样，这种特别的表达方法在C，C++以及现在的Java中顽固地保留下来了。所以倘若您习惯将e作为自然对数的基数使用，那么在Java中看到象“1.39e-47f”这样的表达式时，请转换您的思维，从程序设计的角度思考它；它真正的含义是“1.39×10的-47次方”。 ①：John Kirkham这样写道：“我最早于1962年在一部IBM 1620机器上使用FORTRAN II。那时——包括60年代以及70年代的早期，FORTRAN一直都是使用大写字母。之所以会出现这一情况，可能是由于早期的输入设备大多是老式电传打字机，使用5位Baudot码，那种码并不具备小写能力。乘幂表达式中的‘E’也肯定是大写的，所以不会与自然对数的基数‘e’发生冲突，后者必然是小写的。‘E’这个字母的含义其实很简单，就是‘Exponential’的意思，即‘指数’或‘幂数’，代表计算系统的基数——一般都是10。当时，八进制也在程序员中广泛使用。尽管我自己未看到它的使用，但假若我在乘幂表达式中看到一个八进制数字，就会把它认作Base 8。我记得第一次看到用小写‘e’表示指数是在70年代末期。我当时也觉得它极易产生混淆。所以说，这个问题完全是自己‘潜入’FORTRAN里去的，并非一开始就有。如果你真的想使用自然对数的基数，实际有现成的函数可供利用，但它们都是大写的。” 注意如果编译器能够正确地识别类型，就不必使用尾随字符。对于下述语句： long n3 = 200; 它并不存在含混不清的地方，所以200后面的一个L大可省去。然而，对于下述语句： float f4 = 1e-47f; //10的幂数编译器通常会将指数作为双精度数（double）处理，所以假如没有这个尾随的f，就会收到一条出错提示，告诉我们须用一个“造型”将double转换成float。 2. 转型大家会发现假若对主数据类型执行任何算术或按位运算，只要它们“比int小”（即char，byte或者short），那么在正式执行运算之前，那些值会自动转换成int。这样一来，最终生成的值就是int类型。所以只要把一个值赋回较小的类型，就必须使用“造型”。此外，由于是将值赋回给较小的类型，所以可能出现信息丢失的情况）。通常，表达式中最大的数据类型是决定了表达式最终结果大小的那个类型。若将一个float值与一个double值相乘，结果就是double；如将一个int和一个long值相加，则结果为long。 3.1.14 Java没有“sizeof” 在C和C++中，sizeof()运算符能满足我们的一项特殊需要：获知为数据项目分配的字符数量。在C和C++中，size()最常见的一种应用就是“移植”。不同的数据在不同的机器上可能有不同的大小，所以在进行一些对大小敏感的运算时，程序员必须对那些类型有多大做到心中有数。例如，一台计算机可用32位来保存整数，而另一台只用16位保存。显然，在第一台机器中，程序可保存更大的值。正如您可能已经想到的那样，移植是令C和C++程序员颇为头痛的一个问题。 Java不需要sizeof()运算符来满足这方面的需要，因为所有数据类型在所有机器的大小都是相同的。我们不必考虑移植问题——Java本身就是一种“与平台无关”的语言。 3.1.15 复习计算顺序在我举办的一次培训班中，有人抱怨运算符的优先顺序太难记了。一名学生推荐用一句话来帮助记忆：“Ulcer Addicts Really Like C A lot”，即“溃疡患者特别喜欢（维生素）C”。助记词运算符类型运算符 Ulcer Unary + - ++ – [[ rest...]] Addicts Arithmetic (and shift) * / % + - << >> Really Relational > < >= <= == != Like Logical (and bitwise) && & ^ C Conditional (ternary) A > B ? X : Y A Lot Assignment = (and compound assignment like =) 当然，对于移位和按位运算符，上表并不是完美的助记方法；但对于其他运算来说，它确实很管用。 3.1.16 运算符总结下面这个例子向大家展示了如何随同特定的运算符使用主数据类型。从根本上说，它是同一个例子反反复复地执行，只是使用了不同的主数据类型。文件编译时不会报错，因为那些会导致错误的行已用//!变成了注释内容。 //: AllOps.java // Tests all the operators on all the // primitive data types to show which // ones are accepted by the Java compiler. class AllOps { // To accept the results of a boolean test: void f(boolean b) {} void boolTest(boolean x, boolean y) { // Arithmetic operators: //! x = x y; //! x = x / y; //! x = x % y; //! x = x + y; //! x = x - y; //! x++; //! x--; //! x = +y; //! x = -y; // Relational and logical: //! f(x > y); //! f(x >= y); //! f(x < y); //! f(x <= y); f(x == y); f(x != y); f(!y); x = x && y; x = x \|\| y; // Bitwise operators: //! x = ~y; x = x & y; x = x \| y; x = x ^ y; //! x = x << 1; //! x = x >> 1; //! x = x >>> 1; // Compound assignment: //! x += y; //! x -= y; //! x = y; //! x /= y; //! x %= y; //! x <<= 1; //! x >>= 1; //! x >>>= 1; x &= y; x ^= y; x \|= y; // Casting: //! char c = (char)x; //! byte B = (byte)x; //! short s = (short)x; //! int i = (int)x; //! long l = (long)x; //! float f = (float)x; //! double d = (double)x; } void charTest(char x, char y) { // Arithmetic operators: x = (char)(x y); x = (char)(x / y); x = (char)(x % y); x = (char)(x + y); x = (char)(x - y); x++; x--; x = (char)+y; x = (char)-y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: x= (char)~y; x = (char)(x & y); x = (char)(x \| y); x = (char)(x ^ y); x = (char)(x << 1); x = (char)(x >> 1); x = (char)(x >>> 1); // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; x <<= 1; x >>= 1; x >>>= 1; x &= y; x ^= y; x \|= y; // Casting: //! boolean b = (boolean)x; byte B = (byte)x; short s = (short)x; int i = (int)x; long l = (long)x; float f = (float)x; double d = (double)x; } void byteTest(byte x, byte y) { // Arithmetic operators: x = (byte)(x y); x = (byte)(x / y); x = (byte)(x % y); x = (byte)(x + y); x = (byte)(x - y); x++; x--; x = (byte)+ y; x = (byte)- y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: x = (byte)~y; x = (byte)(x & y); x = (byte)(x \| y); x = (byte)(x ^ y); x = (byte)(x << 1); x = (byte)(x >> 1); x = (byte)(x >>> 1); // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; x <<= 1; x >>= 1; x >>>= 1; x &= y; x ^= y; x \|= y; // Casting: //! boolean b = (boolean)x; char c = (char)x; short s = (short)x; int i = (int)x; long l = (long)x; float f = (float)x; double d = (double)x; } void shortTest(short x, short y) { // Arithmetic operators: x = (short)(x y); x = (short)(x / y); x = (short)(x % y); x = (short)(x + y); x = (short)(x - y); x++; x--; x = (short)+y; x = (short)-y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: x = (short)~y; x = (short)(x & y); x = (short)(x \| y); x = (short)(x ^ y); x = (short)(x << 1); x = (short)(x >> 1); x = (short)(x >>> 1); // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; x <<= 1; x >>= 1; x >>>= 1; x &= y; x ^= y; x \|= y; // Casting: //! boolean b = (boolean)x; char c = (char)x; byte B = (byte)x; int i = (int)x; long l = (long)x; float f = (float)x; double d = (double)x; } void intTest(int x, int y) { // Arithmetic operators: x = x y; x = x / y; x = x % y; x = x + y; x = x - y; x++; x--; x = +y; x = -y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: x = ~y; x = x & y; x = x \| y; x = x ^ y; x = x << 1; x = x >> 1; x = x >>> 1; // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; x <<= 1; x >>= 1; x >>>= 1; x &= y; x ^= y; x \|= y; // Casting: //! boolean b = (boolean)x; char c = (char)x; byte B = (byte)x; short s = (short)x; long l = (long)x; float f = (float)x; double d = (double)x; } void longTest(long x, long y) { // Arithmetic operators: x = x y; x = x / y; x = x % y; x = x + y; x = x - y; x++; x--; x = +y; x = -y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: x = ~y; x = x & y; x = x \| y; x = x ^ y; x = x << 1; x = x >> 1; x = x >>> 1; // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; x <<= 1; x >>= 1; x >>>= 1; x &= y; x ^= y; x \|= y; // Casting: //! boolean b = (boolean)x; char c = (char)x; byte B = (byte)x; short s = (short)x; int i = (int)x; float f = (float)x; double d = (double)x; } void floatTest(float x, float y) { // Arithmetic operators: x = x y; x = x / y; x = x % y; x = x + y; x = x - y; x++; x--; x = +y; x = -y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: //! x = ~y; //! x = x & y; //! x = x \| y; //! x = x ^ y; //! x = x << 1; //! x = x >> 1; //! x = x >>> 1; // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; //! x <<= 1; //! x >>= 1; //! x >>>= 1; //! x &= y; //! x ^= y; //! x \|= y; // Casting: //! boolean b = (boolean)x; char c = (char)x; byte B = (byte)x; short s = (short)x; int i = (int)x; long l = (long)x; double d = (double)x; } void doubleTest(double x, double y) { // Arithmetic operators: x = x y; x = x / y; x = x % y; x = x + y; x = x - y; x++; x--; x = +y; x = -y; // Relational and logical: f(x > y); f(x >= y); f(x < y); f(x <= y); f(x == y); f(x != y); //! f(!x); //! f(x && y); //! f(x \|\| y); // Bitwise operators: //! x = ~y; //! x = x & y; //! x = x \| y; //! x = x ^ y; //! x = x << 1; //! x = x >> 1; //! x = x >>> 1; // Compound assignment: x += y; x -= y; x = y; x /= y; x %= y; //! x <<= 1; //! x >>= 1; //! x >>>= 1; //! x &= y; //! x ^= y; //! x \|= y; // Casting: //! boolean b = (boolean)x; char c = (char)x; byte B = (byte)x; short s = (short)x; int i = (int)x; long l = (long)x; float f = (float)x; } } ///:~ 注意布尔值（boolean）的能力非常有限。我们只能为其赋予true和false值。而且可测试它为真还是为假，但不可为它们再添加布尔值，或进行其他其他任何类型运算。在char，byte和short中，我们可看到算术运算符的“转型”效果。对这些类型的任何一个进行算术运算，都会获得一个int结果。必须将其明确“造型”回原来的类型（缩小转换会造成信息的丢失），以便将值赋回那个类型。但对于int值，却不必进行造型处理，因为所有数据都已经属于int类型。然而，不要放松警惕，认为一切事情都是安全的。如果对两个足够大的int值执行乘法运算，结果值就会溢出。下面这个例子向大家展示了这一点： //: Overflow.java // Surprise! Java lets you overflow. public class Overflow { public static void main(String[] args) { int big = 0x7fffffff; // max int value prt("big = " + big); int bigger = big 4; prt("bigger = " + bigger); } static void prt(String s) { System.out.println(s); } } ///:~ 输出结果如下： big = 2147483647 bigger = -4 而且不会从编译器那里收到出错提示，运行时也不会出现异常反应。爪哇咖啡（Java）确实是很好的东西，但却没有“那么”好！对于char，byte或者short，混合赋值并不需要造型。即使它们执行转型操作，也会获得与直接算术运算相同的结果。而在另一方面，将造型略去可使代码显得更加简练。大家可以看到，除boolean以外，任何一种主类型都可通过造型变为其他主类型。同样地，当造型成一种较小的类型时，必须留意“缩小转换”的后果。否则会在造型过程中不知不觉地丢失信息。 Copyright © quanke.name 2016 all right reserved，powered by Gitbook该文件修订时间： 2018-03-13 01:23:10 results matching "" No results matching ""
Hearing what one has typed using a computer brings undescribable feeling, especially to armature computer operators. This is the reason why the audio books are getting popular now a days. In this article i will show how you can convert text into audio using the Notepad. Why Use Notepad? Because it is the most commonly used text editor everyone uses. That’s the reason. HOW TO CONVERT TEXT INTO AUDIO USING NOTEPAD STEP 1: Open Notepad. You can do this by searching for Notepad in the windows search Or Press Win+R and type Notepad and Press Enter. STEP 2: Now, Copy the following program and paste it in Notepad: Dim message, sapi message = InputBox(“A Best Text to Audio converter”+vbcrlf+”From – http://www.allusefulinfo.com”,”Text to Audio converter”) Set sapi = CreateObject(“sapi.spvoice”) sapi.Speak message STEP 3: Now lets save the code we entered above. The code has to be saved in .vbs format rather than standard .txt format. STEP 4: Now navigate yourself to the location where the code is stored. in my case it was stored on the desktop so i will just go to the desktop and check for it. You can see that the file we saved was in the Visual Studio Script (vbs) format. STEP 5: Now just double click on the saved file and Enter the text you want to hear in the text box. The moment you click OK , You can hear the text you entered as Audio. Advertisements
How To Measure Exhaust Pipe? A Complete Guide: Poor performance of a vehicle can be the result of installing an incorrect exhaust pipe. Now, how would you find the correct size of exhaust pipe only specified for your vehicle? So, how to measure exhaust pipes? Well, you need to follow a simple method by using a measuring tape. Measure the outside diameter of the pipe if the pipe has a defined circle. Or else, take the help of a string to measure the size of the pipe. Lastly, match the obtained size with the closest size for exhaust pipes. A correct exhaust pipe improves fuel efficiency and functionality of the engine. To help you out, the precise measuring method for the exhaust pipe is included below. Let’s dive into it! What Does An Exhaust Pipe Do? The exhaust pipe is the part of the exhaust system that carries harmful gas and fumes out of the vehicles’ engines and cabins. Generally, the exhaust system gathers exhaust gasses from the cylinders. Then, the collected gas passes through the exhaust pipe while being converted into a less toxic end product. Apart from making the expelled gasses less toxic, the pipe also ensures to emit the harmful gasses that can cause damage to the engine. The longer the gasses stay in the exhaust pipe, the more it decreases the performance of the vehicle. But, an efficient and the right exhaust pipe helps to remove the toxic gasses as soon as possible from the vehicle. Thus, it allows fresh oxygen that results in increasing the performance of the vehicle. Faster oxygen intake results in making complete use of the fuel. Meanwhile, complete utilization of the fuel also contributes to increasing fuel efficiency. Furthermore, maximizing adequate fuel usage is facilitated by the quick departure of gasses through the exhaust pipe. However, it also contributes to reducing the noise when the gas passes through the exhaust pipe. Standard Sizes Of An Exhaust Pipe The standard sizes of exhaust pipe for different engine size and exhaust system is tabulated below: Engine size (CI) Horsepower (HP) Single exhaust system Double exhaust system 150-200 100 2” 2”-2 ¼“ 150 2 ¼” 2”-2 ¼“ 200 2 ½” 2”-2 ¼“ 200-250 150 2 ¼“ 2”-2 ¼“ 200 2 ½” 2”-2 ¼“ 250 2 ½” 2”-2 ¼“ 250-300 200 2 ½” 2 ¼ “ 250 2 ½” 2 ½” 300 3” 2 ½” 300-350 250 3” 2 ½” 300 3” 2 ½” 350 3½” 2 ½” 350-400 300 3” 2 ½” 350 3½” 2 ½” 400 3½” 2 ½” 400-450 350 3½” 2 ½” 400 4” 3” 450 4” 3” 450-500 400 4 ½” 3 ½” 450 4 ½” 3 ½” 500 4 ½” 3 ½” Things You Should Know Before Measuring An Exhaust Pipe The size of the exhaust pipe is specified for different vehicles. Still, the owner can upgrade the size of the pipe according to the need. However, the upgraded pipe might not always work effectively for the vehicle. For choosing the correct exhaust pipe for the vehicle, you should consider the following things. Engine Size: Engine size is the first thing to consider before measuring an exhaust pipe. According to the engine size, the vehicle requires a specified size of pipe. For example, larger engines give off a larger amount of mixture of gasses. This results in a larger amount of gas emission. Consequently, it requires a large exhaust pipe. Moreover, installing an incorrect-sized pipe would affect the flow of gas through the exhaust pipe. And the overall performance of the engine would also get disrupted. That’s why using the correct size exhaust pipe is always recommended. Performance: The performance of the vehicle is highly affected by the emission of gas from the vehicle. Well, when the fuel gets utilized, the cylinders release the gas. And the faster the gas travels outside the vehicle, the better the performance of the car will be. That’s why many recommend installing larger-sized exhaust pipes. But, it will also add cost and facilitate other problems, which are also described in the below section. Instead, measuring the exact size of the exhaust pipe should be given priority. How To Measure Exhaust Pipe Properly Sound Preference: The exhaust pipe also influences the noise in the vehicle. Large-sized pipes tend to give off more sounds than smaller ones. Well, the sound is also affected by the resonance system mainly. All the associated parts will also require to be changed due to changing the pipe size. Single And Double Exhaust Systems: The single exhaust system only has one pipeline moving toward the outside. On the other hand, a double exhaust system contains two pipelines to release the gas. Two pipelines work more efficiently by increasing the flow of gas and removing the gas faster. However, not all cars facilitate the double exhaust system. However, the size of the exhaust pipe would also vary with the exhaust system that the car comes with. Associated Parts: The exhaust system consists of the exhaust manifold, oxygen sensor, catalytic converter, and exhaust muffler. Well, all these parts collectively work to get the gas out of the vehicle. The exhaust manifold collects the gas and moves the gas through the exhaust pipe. In the pathway, the gasses get reduced to less toxic components. All these parts come in a standard size for each car. That’s why the exhaust pipe should be installed while considering the associated parts and specifications of the vehicle. How To Measure Exhaust Pipe Properly? Correct measurement of the exhaust pipe is required to ensure the increasing performance of the vehicle. The thickness of the exhaust pipe is generally kept between 1.5-1.6 mm. So, measuring the size does not require measuring the thickness. Any pipes in the marketplace will come with this desired thickness. Furthermore, you can follow the below steps to measure the size of the exhaust pipe. Step 1: Measure The Outside Diameter Generally, the outside diameter (OD) of the exhaust pipe is taken as the size of the pipe. If the pipeline stands out, you can easily measure the end of the pipe and the diameter. Especially when the pipe has a defined circular end with no alteration. To measure the diameter, take a measuring tape or ruler. • Now, hold the tape or ruler from one end of the outside to the other end of the outside of the pipe. • Measure the pipe while keeping the circular center in the middle and holding the tape straight from up to down. • Mark the measurement and note it down. This is the observed diameter of the exhaust pipe. Now, match it with the nearest available pipe size to get the correct size. Step 2: Measure The Circumference If the pipe does not have a perfectly circular shape, you need to measure the pipe from the center. To measure the middle circular diameter of the pipe, you need to find the circumference. To measure the circumference, you need a rope or string. Remember, the string should not be elastic. Now, hold the rope around the pipe and measure the circumference. Do not stretch the rope. Just hold it, not too tight or too softly. Lately, mark the measurement. Then, put it on the ruler to get the correct measurement of the circumference. Afterward, divide the circumference by pi, which is 3.1416, to get the diameter. This is the observed diameter of the exhaust pipe. You should now match the exhaust pipe size with the nearest pipe size. For example, if your obtained diameter of the pipe is 2.60”, what should be the size of the pipe? In that case, the nearest available pipe size is 2.50” and 2.75”. Well, you should take the pipe of 2.75” as the other one might become small for the vehicle. The nearest pipe size is, generally, the correct size of the pipe. Comparing the sizes is important as you might not get a round figure. Necessity Of Perfect Measurement Of Exhaust Pipe Exhaust pipe measurement is required to install it in the vehicle efficiently. Well, the exhaust pipe should be chosen according to the specifications of the car engine. The main function of the exhaust pipe is to expel the gasses as fast as possible without any hindrance. Engine size highly influences the size of the exhaust pipe. As the amount of gas discharged from cylinders also influences their flow through the exhaust pipe. Moreover, putting a large size exhaust would not help as the amount of expelled gas would not be changed. Rather, it will put the engine under pressure to work hard. Also, it can create too much back pressure that can hamper the power production of the engine. It can also reduce fuel efficiency. Associate parts with the exhaust system, such as resonance and muffler size, will also need to change following the exhaust pipe. So buying the larger pipe will only cost money. On the other hand, choosing smaller-sized pipes can be an obstacle to the flow of gas. As the steady flow gets hampered, it gives load to the exhaust system. The excess pressure can also damage the pipe. And does not let the gas out of the pipe efficiently. That’s why choosing the optimum pipe size according to the manufacturer’s specifications should be given priority. Conclusion Overall, how to measure exhaust pipes seems easier as long as you know the correct method. You can measure the size by measuring the diameter directly of a circular pipe. Or else you can determine the circumference first and then find the value of the diameter. In both ways, you can get the correct diameter for the pipe. However, you can also upgrade the pipe size if required. Remember to choose the correct pipe size for the vehicle to get the best performance of the car. Michael is the owner of Michael's Plumbing. He has experience over 15 years solved thousands of plumbing issues. 100% customer satisfaction made him best in this sector. Finally he decided to share his skills, experience and techniques through this PipesYard blog. Hopefully each and every post of this blog will be helpful for people seeking piping help. Leave a Comment
Short introduction to Reactive Extensions The Reactive Extensions (Rx) is a library for programming with asynchronous data streams. It was originally developed at Microsoft for the .NET Framework but is now available as a seperate library. Meanwhile it has been ported to many other languages and platforms as e.g. Java, Scala, C++, Clojure, JavaScript, Python, Groovy, JRuby, and others. Rx can be used in desktop as well as web-based applications on either server or client side. A few common situations / problems where you might benefit from using Rx are: • Unresponsive UI • Long running computations that could potentially block other threads • Asynchronous programming where multiple sources of data and events have to be combined • Enabling and managing concurrency on backend independent of the consumers implementation • Handling errors and cancellation of asynchronous tasks In this post I will introduce some general ideas behind Rx and show how it can help in some of the situations stated above. If you are new to Rx it will give you a basic understanding of what Rx is and where it can be useful to apply. The examples are all presented in C#. But the concepts apply to all programming languages. Definition A definition of Rx could be: Rx is a library for programming with asynchronous data streams. It is a combination of the observer and the iterator pattern as described by the Gang of Four combined with ideas and concepts from functional programming. Querying Data To get an idea when it is useful to use Rx let us look at ways how to retrieve data. Basically a query result can be either a single object or a collection of objects. The data can be queried either synchronous or asynchronous. This produces the following matrix: SingleMultiple Sync Async Sync Single A function that returns a single object and is executed synchronously is probably one of the most common scenarios that we encounter. In the example the function is called GetData() and the return value is of the generic type T as shown in the following table. SingleMultiple SyncT GetData() Async Calling the method and processing the result might look something like this: string s = GetData(); if(s.Equals(x)) // do something else // do something else If this code was implemented behind a user interface the process will be blocked for however long it will take to retrieve the value because this is done synchronously. Sync Multiple When we examine the synchronous query of multiple values we will see the same behaviour except that the return value is a collection of objects of type T. SingleMultiple SyncT GetData()IEnumerable<T> GetData() Async The LINQ library provides a rich toolbox of query operators to create, transform, combine and filter collections. In the example however we want to iterate over all the values and do some processing with each of them. IEnumerable<string> values = GetData(); foreach(string value in values) { if(value.Equals(x)) // do something else // do something else } Again this is done synchronously and the UI will be blocked. Async Single To make the UI more responsive values can be queried asynchronously. In case of a single value the function might return a Task<T>. A Task in C# can be seen as the equivalent of the Future in Java. It is the result of an asynchronous computation. SingleMultiple SyncT GetData()IEnumerable<T> GetData() AsyncTask<T> GetData() Processing the result of a Task can be done synchronously which again will block. Task<T> task = GetData(); if (task.Result.Equals(x)) // do something else // do something else So that is not really an improvement. A better workflow when creating a responsive UI would be: • Respond to some user action • Do work on a background thread • Pass the result back to the UI thread • Update the UI To achieve this we should consider the async features in .NET 4.5 or apply a callback. A callback is a function that is passed as an argument and will be called at a later time e.g. with the result of a long running computation. In this case we can use the ContinueWith method of Task that takes a function that is executed asynchronously. task.ContinueWith(t => { if (t.Result.Equals(x)) // do something else // do something else }); Async Multiple Asynchronous computations that yield collections of values can also be handled with callbacks. Actually this is what is done in many cases. But we might quite easily run into several problems as the application becomes more and more complex. E.g. there is no way to start iterating over the result and process the values before the whole computation completes. If loading the whole collection takes a long time it would be very convenient to start working with the first part of the list while the rest is still processing. But when using callbacks this is not possible. And what if we want to process each element again asynchronously and then each result of that yet again… and so on? The consequence would be a deeply nested composition of callbacks which will become very unwieldy and won’t be easily maintainable any more. With nested composition callbacks become unwieldy and lead to the need to synchronize. This is where Rx comes into play with the main type IObservable<T> – the counter part of IEnumerable<T>. SingleMultiple SyncT GetData()IEnumerable<T> GetData() AsyncTask<T> GetData()IObservable<T> GetData() IObservable<T> represents an asynchronous data stream of objects of type T. Consuming the result is quite simple: IObservable<T> o = GetData(); o.Subscribe(t => { if (t.Equals(x)) // do something else // do something else }); The Subscribe method has several overloads. In the example above it takes an action as a single argument which will be applied to each value from the Observable. Composable functions As well as LINQ Rx also includes a rich library of higher-order functions that allow us to transform, combine, filter etc. observable sequences. Those functions are easily composable. This is one of the really great benefits of Rx because it makes querying, combining and transforming asynchronous data streams as simple as applying LINQ queries. Roughly the set of functions can be divided into the following categories: Transform: Select, SelectMany, Aggregate … Filter: Skip, Take, TakeWhile, Where … Combine: Concat, Merge, Zip … Boolean Operators: Any, All, Contains … Mathematical Operators: Max, Min, Average, Count, Sum … Concurrency: ObserveOn, SubscribeOn … Error Handling: OnErrorReturn, OnErrorResume … Here is a simple example of how some of these functions can be combined: IObservable<int> observable = GetData(); observable .Skip(1) .Take(3) .Where(x => x < 10) .Select(x => x + 1) .Subscribe(Console.WriteLine); IEnumerable vs. IObservable To get an even better understanding of what an IObservable<T> is we will compare it to the IEnumerable<T>. IEnumerable The method IEnumerable.GetEnumerator() returns an object of type IEnumerator which iterates over collections and has the following methods: • bool MoveNext() – Advances the enumerator to the next element of the collection. Returns true if it succeeds and false if it fails (when the enumerator points to the last element of the collection or empty list). • T Current{ get; } – Returns the current element in the list. • throws Exception – This is actually not a method of the interface. But an exception might be thrown when calling Current so an exception is a potential implicit return value of Current. In other words the return value of Current could be described as Either<T, Exception> or Tuple<T, Exception> or similar types. • void Dispose() – Disposes of all resources used by the enumerator. The general task of an enumerator is to pull data from a collection. Below we can see the enumerator in action (Note that this is just for demonstration and is not good code. Usually we would use a foreach statement to do the iteration over a collection.): var enumerator = new List<int> { 1, 2, 3 }.GetEnumerator(); while (enumerator.MoveNext()) { Console.WriteLine(enumerator.Current); } enumerator.Dispose(); IObservable Each method of the IEnumerator interface has its corresponding method in the IObserver interface: • OnCompleted() – Notifies the observer that the provider has finished sending data. • OnNext(T) – Pushes the next value to the observer. • OnError(Exception) – Notifies the observer that an error occurred. The general task of an observer is to push data into a stream. Here is an example of how the methods described above can be called in order to create an observable sequence: public static IObservable<int> GetData() { return Observable.Create<int>(o => { o.OnNext(1); o.OnNext(2); o.OnNext(3); o.OnCompleted(); return Disposable.Empty; }); } For detailed information on how this works and on all the query operators and more check out the amazing online resource about Rx .NET by Lee Campbell. The methods of IEnumerator and IObservable relate to each other in a special way as shown in the following diagram. Actually they are dual to each other. IEnumerableIObservable pullpush bool MoveNext()void OnCompleted() T Current { get; }void OnNext() throws Exceptionvoid OnError(Exception) On an observable sequence and on an enumerable sequence the same or at least very similar functions can be applied: // IObservable<string> // that emits 75 Strings GetData() .Skip(10) .Take(5) .Select(x => x + "_transformed") .Subscribe(x => Console.WriteLine("next => " + x)); // IEnumerablee<string> // that contains 75 Strings GetData() .Skip(10) .Take(5) .Select(x => x + "_transformed").ToList() .ForEach(x => Console.WriteLine("next => " + x)); Summary We have seen how Rx can help to make the UI responsive when dealing with computationally expensive tasks. Also we could get an idea of how easy and comfortable it is to handle asynchronous data from multiple sources with the use of the LINQ-style query operators. Enabling and managing concurrency on the backend as well as handling exceptions, cancellation etc. are other areas where Rx really shines. But these details go beyond the scope of this post. For more information on that please check out the resources below. Things to keep in mind: • Rx is a library for programming with asynchronous data streams • Rx provides a rich library of composable functions to transform, combine, filter etc. • The key types are IObserver<T> and IObservable<T> • The pair of types IObservable<T> and IObserver<T> is dual to the pair IEnumerable<T> and IEnumerator<T> • Rx provides rich functionalities for a scenario where a producer asynchronously pushes data to a consumer Resources The header image “Long exposure head lamps & tail lights” by JC+A is licensed under a Creative Commons Attribution 2.0. The picture was modified to fit this article. Weitere interessante Artikel
LinuxQuestions.org LinuxQuestions.org (/questions/) - Slackware (http://www.linuxquestions.org/questions/slackware-14/) - - Slackware 10.1 how to go to the testing/2.6 kernel and back again? (http://www.linuxquestions.org/questions/slackware-14/slackware-10-1-how-to-go-to-the-testing-2-6-kernel-and-back-again-514364/) bekker 12-28-2006 01:20 PM Slackware 10.1 how to go to the testing/2.6 kernel and back again? Dear all, I'm running a slackware 10.1 with the 2.4 IDE kernel and LVM1. Root is on a normal EXT3 partition. I want to setup an encrypted file system (not root) with truecrypt. However truecrypt requires a 2.6 kernel. So I installed the 2.6.10 kernel from /testing. I then found out that LVM1 does not work with the 2.6 kernels. My question is: when I install the devicemapper and LVM2 packages (after removing LVM1), will I be able to return to my old 2.4 kernel with LVM1 by just removing LVM2 and re-installing LVM1? Will activating the PV/LV's with LVM2 change them, to make them incompatible with LVM1? I have a few custom written programs running on this system so I try to modify the 10.1 defaults as little as I can. Thanks for your time. Chera bsdunix 12-29-2006 01:28 PM Quote: My question is: when I install the devicemapper and LVM2 packages (after removing LVM1), will I be able to return to my old 2.4 kernel with LVM1 by just removing LVM2 and re-installing LVM1? Researching the "LVM HOWTO" in answer to FAQ question 4.1.7 I found this: "... Be warned that it's not always possible to revert back to lvm 1 format metadata." http://www.tldp.org/HOWTO/LVM-HOWTO/lvm2faq.html#AEN348 I suggest you post your question to the LVM mailing list to get a more definative answer. http://www.tldp.org/HOWTO/LVM-HOWTO/maillists.html This is a case where system backups will come in real handy! :) Good Luck. bekker 12-30-2006 09:45 AM BSDunix, thanks for your reply. The same documents states that "4.1.6. Does LVM 2 support VGs and LVs created with LVM 1? Yes. LVM 2 will activate and operate on VG and LVs created with LVM 1. The exception to this is snapshots created with LVM 1 - these should be removed before upgrading. Snapshots that remain after upgrading will have to be removed before their origins can be activated by LVM 2. " That seems to imply that you can run LVM1 VG/LV's with LVM2 without converting them to LVM2 format, as long as you have no snapshots, which I don't have. Also the quick start for going from LVM1 to LVM2 does not say that you need to use vgconvert to update LVM1 volumes to the LVM2 format. regards Chera All times are GMT -5. The time now is 07:34 PM.
dx12 crash Forum discussion tagged with dx12 crash. 1. Avaritia Question Whenever I play a game that uses DX12 it crashes after 10-20 min what should I do to fix it? So I recently (a week ago) got a new prebuilt desktop, since it was cheaper compared to going DIY in my country, and for some reason whenever I play a game that uses DX12 it would crash after 10 to 20 min. I know its DX12 since whenever I play a game that uses DX11 or has the option to choose... 2. G Question Getting desperate with new build. Can't play use any kind of recording software such as Instant Capture on games that use DX12 without crashing. It has been about a month now since I built my new PC, and buy new I mean new everything except for my GPU. Here are my specs below. Windows 11 Pro Gigabyte X670 Elite AX (using latest BIOS F7b) Ryzen 7 7700X MSI 3080ti Gaming X (same GPU from previous build) G.Skill 32GB RAM 6000MHz EXPO... 3. RobinGames Question PC Shuts down and Restarts when playing DX12 Games or running 3D mark fire strike test PC Specs: X570 Taichi Motherboard 4x8 Corsair Vengeance RGB 3200mhz ram CL16 5600x Ryzen CPU 6800xt Merc 319 XFX RM850X Corsair Power Supply (1 1/2 years old) 2TB Sabrent Rocket SSD 4TB Toshiba 300X HDD (2 or 3 years old) 9 fans on a cooler master Mesh TD300 case Alright fellas I have... 4. G [SOLVED] Computer crashes an hour into gaming but only for DX12 Got an interesting issue or at least I think so. Hopefully this is the appropriate thread, let me know if not. Have been troubleshooting for a month, have made progress but now feel like I’m chasing a red herring maybe. So I had built a fairly new PC, Used some Parts from my previous build...
ابحث فى المدونة أحدث المواضيع الطرق المختلفه لبدء تشغيل المحرك الحثي Induction Motor Different Ways For Induction Motor Starting من المعروف ان في بدايه تشغيل المحرك Induction Motor يجب أن يتحمل مرور تيار عالي للتغلب على القصور الذاتي للكتلة الميكانيكية مما يستلزم عزم Torque كبير فى البداية ، ويبلغ التيار فى بداية التشغيل حوالي 6 مرات قدر تيار التحميل ، وقد يسبب التيار العالي هبوطا فى الجهد الكهربي المغذي للشبكة مما يؤدي إلى فصل بعض الأحمال ، لذلك يجب ألا يزيد الهبوط فى جهد المصدر أثناء بدء تشغيل المحرك عن 4 % ويتطلب ذلك أن يكون جهاز بدء التشغيل قادرًا على تحميل المحرك عدة مرات خلال الساعة الواحدة دون مشاكل للشبكة . لذلك هناك اكثر من طريقه مختلفه تستخدم لبدء تشغيل Induction Motor ومنها :- 1 - Direct on-line starting (DOL) التشغيل المباشر من خط التغذية This is the simplest mode, where the stator is directly connected to the mains supply. The motor starts with its own characteristics. When it is switched on, the motor behaves like a transformer with its secondary, formed by the very low resistance rotor cage, in short circuit. There is a high induced current in the rotor which results in a current peak in the mains supply: Current on starting = 5 to 8 rated Current. The average starting torque is: T on starting = 0.5 to 1.5 rated T. In spite of its advantages (simple equipment, high starting torque, fast start, low cost), direct on-line starting is only suitable when: - the power of the motor is low compared to that of the mains, which limits interference from inrush current, - the machine to drive does not need to speed up gradually or has a damping device to limit the shock of starting, - the starting torque can be high without affecting machine operation or the load that is driven. 2 - Star/Delta بدء التشغيل بإستخدام طريقة نجمة – دلتا This method requires both connections for each phase (six in all) to be taken to the starter. Three contactors are used to first connect the motor in star and then to delta after a given time. Connecting the motor in star reduces the voltage applied to each winding to about 60% of the line voltage. This reduces the starting torque and current (typically 3.5 x FLC). After a given time the motor is switched to delta connection and then runs as if direct-on-line. Its main advantages are that it is relatively simple and low cost. The major problem with this method is that the reduced voltage level is in a single stage and is fixed. sometimes this voltage is not ideal, the torque it produces (65% of full load torque) may be too small and the motor stalls or does not give complete acceleration, or if it is too great the motor still starts with a pronounced snatch. The star/delta transition will produce a second current and torque peak which is almost the equivalent of having two direct-on-line starts. On some loads the motor sometimes almost stalls during this transition time. This method of starting does however have the advantage of being a low cost and simple solution if its limitations can be tolerated . Advantages : 1- Low cost and simple Disadvantages 1- Torque too high – causes snatch 2- Torque too low – motor stalls 3- Transition peak up to 20 x In 4- Motor can stall in transition 3 - Auto Transformer This method uses transformer action to reduce the voltage applied to the motor and current seen by the supply. An improved torque/amp ratio is achieved and starting current is typically 3 x FLC, depending on the voltage rapping selected. Normally the voltage is applied to the motor in voltage steps through the transformer with the taps being selected through contactors. Typical tappings are 50%, 70%, followed by full voltage being applied to the motor. The major disadvantages are size and cost, and of course the mechanical snatch at switch on is not controllable and can still cause problems. Also once the tappings have been selected, it may be necessary to change them according to changes in load parameters. Advantages 1- Simple operation Disadvantages 1- Poor controllability 2- Bulky 3- Very Expensive 4 - Soft Starters The soft start is designed to apply an adjustable voltage to the motor and increase this voltage gradually over a user-selectable acceleration period. The acceleration time being dependent on the application and desired characteristics. The added advantage of this method of reduced voltage control is that the motor can also be stopped gradually by slowly reducing the output voltage to the .‘Soft Stop’ feature offers a smooth stop in many process industries such as pumps, where fast stops can result in ‘water hammer’ and mechanical damage Advantages 1- Reduced starting current 2- Reduced starting torque 3- Less mechanical stress 4- Improved control of acceleration and deceleration 0 comments: data: commentLabel/ Related Posts Plugin for WordPress, Blogger...
Butt Exercise Mini Series – Yoga and Pilates Butt Exercise Mini Series – Yoga and Pilates Yoga Yoga has been called a science or technology of liberation for many years however there is more to this art than just “inner peace” it is a great for developing muscles too! Yoga seeks to provide the student with a practical path towards the common goal of liberation and inner peace As explained above, Yoga is a diverse tradition, which makes it quite difficult to provide a concise summary of the philosophy and art. One approach is to consider common elements that are found in all (or nearly all) branches of the tradition. Without boring you with the history of yoga, the main premise in which yoga works is that you must find inner peace with all you elements and yoga is the practical means in which you can do that. Returning to the practical side, the first step for any yoga student is to find a suitable teacher which is able to put forward his teachings so you can find the inner peace yourself. Traditionally, this relationship would be with a guru who is seen as an embodiment of the Divine who has spent his whole life perfecting his yoga skills. The student’s life would often involve manual service to the guru as a form of payment for instruction. A guru may also found an ashram or order of monks. Many gurus of the past have written modern translations of classical texts, explaining how their particular teachings should be followed and interpreted. In practice, the modern western student is much more likely to attend a local yoga course and receive instruction from a teacher who are themselves practicing the style of a particular school founded by a guru so you won’t actually be learning from a guru, rather you’ll be going through the motions of another student’s interpretation of the guru’s work. For our purposes, yoga in its most basic teaches all the basic movements and techniques such as proper breathing, relaxation and pin-pointing and above all concentration in the form of meditation. Although it isn’t directly related to having a great butt, many of the practices of yoga has the benefit of balancing muscle distribution. Thus, if you are imbalanced in the butt region joining yoga will have the benefit of helping to indirectly re-distribute muscles to areas more in-need and reducing in areas which aren’t needed for a overall better shape. Yoga, as far as I’m concern has the most beneficial side-effects too, it is extremely calming and creates “peace” in the daily hustle and bustle of life. It creates a very proportioned physique and in the process makes you look much more attractive by giving you a neutral butt. Pilates Pilates is a physical fitness system that was developed in the early 20th century by a fitness expert called Joseph Pilates. Pilates called his exercises a form of contrology which refers to the way the method encourages the use of the mind to control the muscles. The program focuses on the core postural muscles that help keep the body balanced and are essential to providing support for the spine and any other outer regions of the body. In particular, Pilates exercises teach awareness of breathing and alignment of the spine, and to strengthen the deep torso muscles, which are important to help build core muscles such as lower back muscles which prevent back pain. Of late, Pilates as grown in popularity as the trend is to have long lean muscles rather than bulky large muscles and this bring us to how it benefits the butt especially when you are trying to lean out the butt muscles, get better definition and even loose some fat in the butt region. The general goals of Pilates are as follows according to Joseph Pilates published books: • Enhances body alignment • Strengthens the abs, back and stabilizer muscles • Stretches, strengthens and relaxes your body • When modified, is gentle enough for pregnant women, yet challenging enough for advanced exercisers. • Incorporates exercises that target your abs, back and legs, making your body strong and flexible. Some may mistake Pilates as a form of Yoga, rather what makes Pilates different from Yoga is that it’s more dynamic. In many moves, you hold your torso in place while moving your limbs in different directions, which challenges your balance, core strength, stability and flexibility. There are a whole host of exercises for the butt which Pilates is great for, however for the most part, I believe them to be sub-standard when it comes to sheer muscle development. Pilates should be used to lengthen and define muscles rather than build them. As an added benefit, the majority of Pilates exercises are very high reps (50-100 reps) which is also very good for burning fat and adding definition. More articles from the Butt Exercise Series: Can A Workout Guide Help Guide You In The Right Direction? You’ve decided to start working out. However, when you look into the best types of exercise to suit your purpose, you’re faced with an overwhelming plethora of choices! Do you join a gym, do aerobics or just start jogging around the block every day? Different types of exercises suit different people and different weight loss / fitness goals. You also need to factor in things like – what type of exercise do you most enjoy doing. Or to put it another way – what type of exercise is going to be sustainable for you in the long term. There is no point deciding to take up running if you hate running or have health issues that would make running unfeasible. Likewise, joining a gym is out if you can’t afford the membership fees or there isn’t one convenient for you. The Walking Workout – Walking Your Way To Better Fitness One of the most convenient, easiest and popular ways to exercise is walking. Walking is an excellent form of exercise and suits pretty much every level of fitness. Additionally, nearly everyone can fit some walk time into their daily routine, even if it’s just walking the dog on a daily basis. It’s relatively easy to find an exercise partner to go walking with you, which provides added incentive to keep doing it. As you get fitter, you can increase the distance and intensity at which you walk too. Aerobics – A Great Fitness Workout Option Another very popular type of exercise, especially for women, is aerobics. In some countries, free or cheap council-run aerobics classes are a popular alternative to joining an aerobics club. If you think aerobics might be something you’d like to try, be sure to get hold of an aerobics for fitness guide. This will show you how the various types of exercises will benefit you and importantly, how to do them correctly. Joining A Gym To Work Out Yet another option is joining a fitness center or a gym. This gives you the option of having a variety of workout programs and equipment available to suit. Most also have trainers who can help you plan an appropriate workout and answer any questions. When choosing a fitness center, there are a couple of things to bear in mind. Good service is essential and well-maintained equipment of paramount importance. Exercise equipment in good condition and maintained correctly doesn’t generally squeak, rattle or make a lot of noise when used! The Ultimate Convenience – Working Out At Home You may even decide to purchase your own exercise equipment to set up at home. That’s the ultimate in convenience! However, it is an expensive outlay so be sure to select the best equipment for losing weight the way you want to lose it. If you don’t plan to do much strength training, don’t go overboard buying a ton of strength building equipment. Choose The Right Exercises With A Workout Guide Regardless of the type of exercise you choose, most workout guides advise you to start out the same way. Begin with a low level of intensity then build up to higher levels of intensity as your strength and fitness improves. If you start out by doing too much too soon you’ll run the risk of injuring yourself, possibly seriously. Also, bear in mind that depending on the type of exercise being done, workout guides written for women may differ from those for men simply because of differences in muscle bulk and strength. 4.5/5 - (6703 votes) About Linda Bell Linda Bell is in the fortunate position of being able to combine doing something she loves with spending a lot more time with those she loves. She and her family occupy an unassuming cottage in the midst of splendid chaos AKA ‘the garden’ in England. Somewhere in that garden lurks the family dog and a feline with a penchant for getting into trouble. Linda loves researching and writing about health and fitness, something that is dear to her heart after some years spent working in a fitness centre overseas.
NewRecords Takes one parameter, QuantityToAdd Description: The specified number of blank records will be added to the Query Objects memory space. Do not confuse this command with the Database command New which is responsible for creating new Records in the Database. Although this is in most cases not useful, for some algorithms it could be necessary. Note that you can use the %Index built in numeric value to seed such blank records with unique information.
Quantcast Tissue Specificity of Human Angiotensin I-Converting Enzyme. Research paper by Olga V OV Kryukova, Victoria E VE Tikhomirova, Elena Z EZ Golukhova, Valery V VV Evdokimov, Gavreel F GF Kalantarov, Ilya N IN Trakht, David E DE Schwartz, Randal O RO Dull, Alexander V AV Gusakov, Igor V IV Uporov, Olga A OA Kost, Sergei M SM Danilov Indexed on: 26 Nov '15Published on: 26 Nov '15Published in: PloS one Abstract Angiotensin-converting enzyme (ACE), which metabolizes many peptides and plays a key role in blood pressure regulation and vascular remodeling, as well as in reproductive functions, is expressed as a type-1 membrane glycoprotein on the surface of endothelial and epithelial cells. ACE also presents as a soluble form in biological fluids, among which seminal fluid being the richest in ACE content - 50-fold more than that in blood.We performed conformational fingerprinting of lung and seminal fluid ACEs using a set of monoclonal antibodies (mAbs) to 17 epitopes of human ACE and determined the effects of potential ACE-binding partners on mAbs binding to these two different ACEs. Patterns of mAbs binding to ACEs from lung and from seminal fluid dramatically differed, which reflects difference in the local conformations of these ACEs, likely due to different patterns of ACE glycosylation in the lung endothelial cells and epithelial cells of epididymis/prostate (source of seminal fluid ACE), confirmed by mass-spectrometry of ACEs tryptic digests.Dramatic differences in the local conformations of seminal fluid and lung ACEs, as well as the effects of ACE-binding partners on mAbs binding to these ACEs, suggest different regulation of ACE functions and shedding from epithelial cells in epididymis and prostate and endothelial cells of lung capillaries. The differences in local conformation of ACE could be the base for the generation of mAbs distingushing tissue-specific ACEs.
Debugging tricky (Arm Cortex-M4) code with register values Sometimes you may run into problems when debugging tricky code. This is the case especially with microcontroller code if you are implementing a bootloader+firmware image solution. Debugger started in bootloader goes haywire and displays garbage or nothing at all when jumping to firmware. Normally one should invest some time in getting debugger symbols aligned properly with source code, but if there are for example some barring deficiencies in environment or debugger UI, one can still figure out a bit more what is going on by using spare register values. Following shows a somewhat obvious, but still possibly helpful technique in brief for aforementioned debugging situations. UPDATE 2021-08-16: To get the job actually done, see my follow-up post But in any case, the rest of the earlier post is here for posterity. In case you are in same kind of situation as me, you probably have found the following screen all too familiar: However, if we set the instruction stepping mode, we can still see the actual assembly code: So, processor is running and it is still reading and running the machine instruction list, if you will. The problem many times becomes, where it actually is and what values is it seeing. Luckily there is a small fix to this, in case one cannot get the debugger to fetch right debug symbols to right offset. We can use individual MCU registers to convoy minimal debug information. The amount available depends on the processor, naturally. I am running myself Cortex-M4 core, from STM32 Nucleo-L432KC evaluation board. Register map (visible while debugging from a tab) looks like this: The actual map is longer, but this is enough for our purpose. As described in the picture, on Cortex-M4 (and many others) we have basically the “normal registers” r0 – r12 available to us. While taking a look at the machine code generated, we can quickly see that registers r0 – r7 are frequently used. Then we have the friend zone r8 – r10 where usually (with varying degrees of usuality) nothing happens. But then we have the r11 and r12 registers. They are very, very, very unlikely to be present in local assembly contexts, so we could in theory pass information via them. But how do we do this? We can inject simple assembly instructions ourselves. Take a look at this MCU startup assembly code: By using simple assembly instructions, like ldr r11, =0xDEB00110; we are able to see where we are at the code. The previous was in pure assembly section. How about actual C functions? The same way, we just use a specific function to lace in our debug info: As we can see, first code line we used was asm("ldr r11, =0xDEB00030"); Next a little bit of suspense. What happens if we start the debugger and enter the “unknown” code one assembly instruction per time? Lets see. After selecting “Instruction Stepping Mode” and register view and then at jump point stepping with F5 one instruction a time, we end up in place unknown to debugger: At this point it is best to ser r11 and r12 to hex display mode: Stepping a bit further reveals a surprise: We found our “custom debug symbol”! This tells us that jumping to the setup section (Reset_Handler) of the “unknown” code worked! Hitting F5 couple of times get us 0xdeb00010. It means that setting the stack pointer was successful. Somewhat important information. Hitting F5 enough gets us to 0xdeb00030. This is great! It means we successfully jumped to the C function SystemInit() showed earliler. Eventually we get to0xdeb00110, which is great information, because it tells us that we successfully jumped back to caller! There is one additional trick to this all. Remember the r12 we have been neglecting? It can be used in parallel with r11 to show information. Here for example we load the address of the location of label __libc_init_array we are going go to and put that information to r12. So when r11 shows 0xdeb000170, we step one assembly instruction further and see that we are going to try to call code at address 0x80018cd: (This is wrong for my use case, because I link stuff beginning to 0x8000000 but flash it to 0x8005000 as I am experimenting with creating bootloader+firmware system where firmware can be anywhere. Now I can inspect the problem and see if I can patch it.) As a general hint, one should use symbols of the form 0xdeb00010, 0xdeb00020, 0xdeb00030 etc, so in case you need to put more info somewhere in between, you still have some numbers left. But wait, there is even more… With a bit of trickery it is possible to break to code position when a register changes. Lets see how it happens. Select debug view, breakpoints, right clcik and select from context menu “Add Watchpoint (C/C++)”: Then write the details. Strangely, in this screen you need both fields; the expression to watch and condition. I used these values: $r11 and $r11 == 0xDEB00110 This signifies to stop when register r11 reads 0xDEB00110 . Next we actually disable the watchpoint because slows down the debugger if continuously running: Next we start the known code and just before jumping to the “unknown code”, we enable the watchpoint: Then we wait a bit, and finally get to the position of our debug identifier! Now we can correspond a bit easier the live assembly code to the code written. As we saw, emitting custom debug symbols via registers can be somewhat useful debug tool if and only if nothing else works. Usually it is best to invest some time to explore how to get the actual debug symbols lined up in the program. Leave a Reply Your email address will not be published. Required fields are marked *
Chronic Diseases health conditions and diseases With regard to end-stage renal illness, Figure four exhibits that the condition is dispersed extensively among numerous areas. The American Cancer Society and other organizations have been trying to make both sufferers and physicians extra conscious and contemplate this analysis if the classic signs are present. Back ache, pelvic ache, bloating, or indigestionThese are common symptoms of daily life, usually related to food intake, muscle spasms or strains, but they also may be seen in ovarian most cancers. Particulate matter, for instance, has been demonstrated to be a probable causal factor in both cardiovascular disease morbidity and mortality. Many studies present a relationship between well being and earnings, with the poorest sections of the population being essentially the most vulnerable. Poor people are at an increased social drawback by way of the incidence of persistent diseases, in addition to access to treatment. They also show decrease charges of acceptance of health-selling behaviours in contrast with other sectors of society. Thus, insurance policies have to favour the poor and appropriately targeted, as poor people are most at risk and have the least energy to impact change. Physical Activity and Nutrition The most necessary change to our healthcare system ever, occurred with the legislating of the Patient Protection and Affordable Care Act (HHS, Healthcare, PPACA, 2010). This regulation has had main impacts, optimistic and negative, on how healthcare is paid for, how healthcare is accessed, and where healthcare could be obtained. The affects have been far reaching and will most likely be the focus of more adjustments to come back in 2017 as a brand new Presidential administration moves into workplace. The U.S. healthcare system continues to expertise phenomenal development of recurring themes of important service supply points throughout the business. As one of the largest employers within the country, and since each citizen needs healthcare at one time or one other, it is important to gain an understanding and stay up to date about the issues that drive our healthcare system (HHS, CMS, 2016). What is advanced care? According to WHO and the CDC, these infectious illnesses are the five most common. Infection occurs whenviruses,micro organism, or other microbesenter your physique and begin to multiply. Telehealth can broaden access to specialty and subspecialty care, in addition to training for rural providers associated to sufferers with persistent situations. RHIhub’s Telehealth Use in Rural Healthcare subject guide has many assets on how telehealth can enhance entry to care in rural communities. For examples of telehealth applications or sources to assist in creating a telehealth program, see RHIhub’s Rural Telehealth Toolkit. Although extra primary analysis could also be needed on some features of the mechanisms that hyperlink diet to health, the presently out there scientific evidence provides a sufficiently robust and believable basis to justify taking action now. Beyond the suitable medical treatment for those already affected, the general public health approach of main prevention is taken into account to be essentially the most cost-efficient, inexpensive and sustainable course of action to cope with the chronic illness epidemic worldwide. Research signifies that staying physically energetic can help forestall or delay sure illnesses, including some cancers, heart illness and diabetes, and likewise relieve melancholy and enhance mood. Check together with your native church buildings or synagogues, senior centers, and purchasing malls for train and strolling packages. Like exercise, your consuming habits are often not good should you live and eat alone. It’s essential for profitable growing older to eat foods rich in nutrients and keep away from the empty calories in candy and sweets. The majority of individuals with asthma are able to manage it with remedy, but nonetheless there is no treatment. Children’s Mercy Allergy, Asthma, and Immunology Telemedicine and Kansas Asthma Initiative are rural program examples that use telemedicine to help rural patients entry subspecialty care. Health-Related Behaviors by Urban-Rural County Classification ”” United States, 2013, a 2017 MMWR article, discovered that smoking will increase with degree of rurality. The report evaluation discovered that solely 74.9% of the noncore or small rural and 76.5% of micropolitan or massive rural residents are nonsmokers, in comparison with eighty one% of the nation as an entire. Regular physical exercise can forestall or delay the development of high blood pressure, and reduces blood strain in persons with hypertension. Individuals affected by persistent illnesses bear a considerable portion of these medical prices.
AI帮你理解科学 AI 生成解读视频 AI抽取解析论文重点内容自动生成视频 pub 生成解读视频 AI 溯源 AI解析本论文相关学术脉络 Master Reading Tree 生成溯源树 Identification of joint stiffness with bandpass filtering ICRA, no. 1 (2001): 2867-2872vol.3 引用69\|浏览27 EI WOS 下载 PDF 全文引用摘要 Proposes a method to identify the joint stiffness of a robot using a bandpass filter. It is based on moving one axis at a time. The dynamic model reduces to a model which is linear in relation to a minimum set of dynamical parameters which have to be identified. These parameters are estimated using the least squares solution of an over de...更多代码数据 0 您的评分 : 暂无评分标签评论数据免责声明页面数据均来自互联网公开来源、合作出版商和通过AI技术自动分析结果，我们不对页面数据的有效性、准确性、正确性、可靠性、完整性和及时性做出任何承诺和保证。若有疑问，可以通过电子邮件方式联系我们：report@aminer.cn
dcsimg March 27, 2015 Hot Topics: Mastering Internet Programming on Mobile Devices: First Steps • November 1, 2004 • By Alex Gusev • Send Email » • More Articles » Working with Sessions and Synchronous Requests An alternative way to get connected is to use InternetConnect-based APIs. The common working flow may be organized as follows: 1. Call InternetConnect to get session handle. 2. Call HttpOpenRequest to define all desired request parameters and request type (HTTP or FTP). 3. Call HttpSendRequest or HttpSendRequestEx to send the request to the remote host. 4. Read the possible answer with InternetReadFile. 5. Close the request handle. 6. Repeat all from Step 2. 7. Close the session handle. In C# terms, all the business looks even simpler: 1. Create an instance of the WebRequest (actually HttpWebRequest) class. 2. Set up all required headers' values if needed. 3. Retrieve the host response as an instance of the HttpWebResponse class. 4. Read data from the response through the Stream and StreamReader classes. To illustrate all that was mentioned above, let's consider a couple of samples in both the C/C++ and C# languages: // C/C++ sample, GET method void CWceHttpDlg::OnButton2() { UpdateData(); BOOL bRC = TRUE; HINTERNET hGETRequest; DWORD dwFlags = INTERNET_FLAG_NO_CACHE_WRITE \| INTERNET_FLAG_KEEP_CONNECTION \| INTERNET_FLAG_IGNORE_CERT_CN_INVALID \| INTERNET_FLAG_IGNORE_CERT_DATE_INVALID\| INTERNET_FLAG_PRAGMA_NOCACHE; LPTSTR pszAcceptTypes [] = {TEXT("text/"), NULL}; TCHAR szServer [1024]; TCHAR szEndpoint [1024]; URL_COMPONENTS crackedURL; int nPort; HINTERNET hConnect; HINTERNET hOpen; CString sHTTPHeader; CString sInfo; if (m_sURL.IsEmpty ()) return; //Crack URL ... ZeroMemory (& crackedURL, sizeof (URL_COMPONENTS)); crackedURL.dwStructSize = sizeof (URL_COMPONENTS); crackedURL.lpszHostName = szServer; crackedURL.dwHostNameLength = 1024; crackedURL.lpszUrlPath = szEndpoint; crackedURL.dwUrlPathLength = 1024; InternetCrackUrl (m_sURL, 0, 0, &crackedURL); nPort = crackedURL.nPort; hOpen = InternetOpen (L"WceHttp", INTERNET_OPEN_TYPE_PRECONFIG, NULL, NULL, 0); if ( !hOpen ) { AfxMessageBox(L"Failed to open WinInet"); return; } hConnect = InternetConnect (hOpen, szServer, nPort, L"", L"", INTERNET_SERVICE_HTTP, 0, 0); if ( !hConnect ) { sInfo.Format(L"InternetConnect failed: %lu", GetLastError ()); AfxMessageBox(sInfo); return; } // Open an HTTP request handle... hGETRequest = HttpOpenRequest (hConnect, L"GET", szEndpoint, NULL, NULL, (LPCTSTR) pszAcceptTypes, dwFlags, 0); if ( !hGETRequest ) { sInfo.Format(L"HttpOpenRequest failed: %lu", GetLastError ()); AfxMessageBox(sInfo); InternetCloseHandle (hConnect); return; } // send the request... sHTTPHeader = L"Content-Type: text/*\r\n"; if (! HttpSendRequest (hGETRequest, (LPCTSTR) sHTTPHeader, sHTTPHeader.GetLength (), NULL, 0)) { sInfo.Format(L"HttpSendRequest failed: %lu", GetLastError ()); AfxMessageBox(sInfo); InternetCloseHandle (hGETRequest); InternetCloseHandle (hConnect); return; } char szBuffer[4096]; DWORD dwNumberOfBytesRead = 0; TCHAR wszTmp[4097]; int i = 0; DWORD dwTotal = 0; while ( InternetReadFile(hGETRequest, szBuffer, 4096, &dwNumberOfBytesRead) && dwNumberOfBytesRead ) { memset(wszTmp,0,sizeof(wszTmp)); MultiByteToWideChar(CP_ACP,0,szBuffer,dwNumberOfBytesRead, wszTmp,sizeof(wszTmp)); i++; dwTotal += dwNumberOfBytesRead; } sInfo.Format(L"Read %d block(s) - %lu byte(s)",i,dwTotal); AfxMessageBox(sInfo); InternetCloseHandle (hGETRequest); InternetCloseHandle (hConnect); InternetCloseHandle (hOpen); } // C# sample, GET method private void cmdConnect_Click(object sender, System.EventArgs e) { string url = txtURL.Text; string proxy = txtProxy.Text; try { if(!"".Equals(txtProxy.Text)) { WebProxy proxyObject = new WebProxy("10.168.2.25:8080",true); proxyObject.Credentials = new NetworkCredential ("alexg", "England2007", "posnet"); // Disable proxy use when the host is local. proxyObject.BypassProxyOnLocal = true; // HTTP requests use this proxy information. GlobalProxySelection.Select = proxyObject; } WebRequest req = WebRequest.Create(url); WebResponse result = req.GetResponse(); Stream ReceiveStream = result.GetResponseStream(); Encoding encode = System.Text.Encoding.GetEncoding("utf-8"); StreamReader sr = new StreamReader( ReceiveStream, encode ); // Read the stream into arrays of 1024 characters Char[] read = new Char[1024]; int count = sr.Read( read, 0, 1024 ); while (count > 0) { String str = new String(read, 0, count); txtOutput.Text += str; count = sr.Read(read, 0, 1024); } } catch(WebException ex) { string message = ex.Message; HttpWebResponse response = (HttpWebResponse)ex.Response; if(null != response) { message = response.StatusDescription; response.Close(); } txtOutput.Text = message; } catch(Exception ex) { txtOutput.Text = ex.Message; } } Both samples synchronously download some file from a Web server by the HTTP "GET" method. If you need to use the "POST" method, the only additional steps you will be required to do are: Win32 API: • Define request content type—"text/xml" or "application/x-www-form-urlencoded". • The request verb should be set to "POST" when calling either the HttpOpenRequest or HttpWebRequest.Method property. • Provide lpOptional and dwOptionalLength parameters to HttpOpenRequest as form data according to the HTTP specification. Sample: HttpSendRequest(hRequest, (LPCTSTR) sHTTPHeader, sHTTPHeader.GetLength (), lpFormData, dwFormDataLength)) C#: • Set the HttpWebRequest.ContentType property to the desired content type. • The HttpWebRequest.Method property should be set to "POST". • Create an instance of the Stream class on the HttpWebRequest object and then call its Write method. Sample: ... // data is an array of length nLen which should be sent WebRequest req = WebRequest.Create(url); req.Method= "POST"; req.ComtentType = "text/xml"; req.ContentLength = nLen; Stream reqStream = req.GetRequestStream(); reqStream.Write(data,0,nLen); reqStream.Close(); WebResponse resp = req.GetResponse)(); ... After such manipulations, data will be sent to the remote server. In turn, the server can return some response that you can interpret as required in your partucular situation. Conclusion In this article, we have discussed the very first and simple tasks of Internet programming on mobile devices. By using the provided information, you are already able to transmit data from/to Web servers. The next articles will overview asynchronous requests, cookies, secure connections, and XML HTTP features. Downloads Download the accompanying C++ code's zip file here. Download the accompanying C# code's zip file here. About the Author Alex Gusev started to play with mainframes at the end of the 1980s, using Pascal and REXX, but soon switched to C/C++ and Java on different platforms. When mobile PDAs seriously rose their heads in the IT market, Alex did it too. Now, he works at an international retail software company as a team leader of the Mobile R department, making programmers' lives in the mobile jungles a little bit simpler. Page 2 of 2 Comment and Contribute (Maximum characters: 1200). You have characters left. Enterprise Development Update Don't miss an article. Subscribe to our newsletter below. Sitemap \| Contact Us Thanks for your registration, follow us on our social networks to keep up-to-date Rocket Fuel
MIL Builder class coremltools.converters.mil.mil.Builder[source] This class is a singleton builder to construct a MIL program. For more information, see Create a MIL program. Importing .ops triggers the installation of all MIL ops into the Builder. For details on each op, see MIL ops. Examples >>> from coremltools.converters.mil.mil import Builder as mb >>> from coremltools.converters.mil.mil import Program, Function >>> prog = Program() >>> func_inputs = {"x": mb.placeholder(shape=[2,3]), >>> "y": mb.placeholder(shape=[2,3])} >>> with Function(func_inputs) as ssa_fun: >>> x, y = ssa_fun.inputs['x'], ssa_fun.inputs['y'] >>> res_var = mb.add(x=x, y=y) # created within ssa_fun block >>> ssa_fun.set_outputs([res_var]) >>> prog.add_function("main", ssa_fun) >>> # Importing ops triggers installation of all ops into Builder. >>> from .ops import defs as _ops static program(input_specs=None, opset_version=None)[source] The mb.program decorator creates a MIL program with a single function (main). The input to main is a tensor. Parameters: input_specs: TensorSpec Describes a tensor. opset_version: AvailableTarget enum Describes the opset version of the program Examples >>> import coremltools as ct >>> @mb.program(input_specs=[mb.TensorSpec(shape=(1,2))], opset_version=ct.target.iOS16) >>> def prog(a): >>> return mb.add(x=a, y=2)
Tag Archives: Minkebe Minkebe National Park Minkebe National Park Life cycle Breeding occurs throughout the year. Depending on the fruiting times of the plants they feed on, activities may shift seasonally. Spread of infectious diseases, notably Ebola Since the early s, outbreaks of Ebola hemorrhagic fever have caused large-scale die-offs of great apes. The animals forage during the day for hours, with peaks of activity in the early morning and late afternoon. Sharing 98% of our genes Bakersfield Sequoia National Park The overall declining trend of both African elephant species calls for increased support by donors, governments of the elephant range states, and the Four subspecies have been identified, based on differences in appearance and distribution: Western chimpanzee P. Physical description The characteristic chimpanzee shape includes arms that extend beyond the knees, opposable thumbs, and a prominent mouth. The skin on the face, ears, palms, and soles of the feet is bare, and the rest of the body is covered with brown to black hair. Young individuals sometimes swing from branch to branch. Social structure Chimpanzees are highly social animals. Their communities consist of loose and flexible groups of males and females fusion-fission societies within a fixed home range, led by a dominant male. Apart from the dominant leader, there are also groups of individuals with some level of authority. Communities of about Minkebe National Park individuals each have been reported in forest, woodland and savanna habitat, but overall size range is around Subgroups may include solitary individuals or diverse groups of both sexes and all ages. These aggregations are temporary and constantly change in composition, regardless of gender and age. Life cycle Breeding occurs throughout the year. Following a gestation period ranging from and days, females give birth to a single young, and occasionally twins, every years. Of these offspring, about three will survive. For the first 6 months, the young is carried around clinging to its mother’s underbelly, and after that it Glacier National Park To Great Falls Mt on its mother’s back. It weans at 3. Although chimpanzees reach sexual maturity at about 7 years, females do not produce offspring until Glacier National Park To Great Falls Mt reach years of age. Chimpanzees may live until they are over Chimpanzees eat with their hands, which they also use to throw objects at enemies and to create tools. Minkebe National Park, they will poke a stick into a termite mound to feed on the insects, and crack nuts open. The animals forage during the day for hours, with peaks of activity in the early morning and late afternoon. Depending on the fruiting times of the plants they feed on, activities may shift seasonally. Chimpanzees sometimes stalk, kill and eat other primates or young antelopes, and may hunt co-operatively. Priority place. Spread of infectious diseases, notably Ebola Yosemite National Park Conditions Following a gestation period ranging from and days, females give birth to a single young, and occasionally twins, every years. Notably, they will poke a stick into a termite mound to feed on the insects, and crack nuts open. Young individuals sometimes swing from branch to branch.
Monarch Butterfly Monarch butterflies are known for the incredible mass migration that brings millions of them to California and Mexico each winter. North American monarchs are the only butterflies that make such a massive journey—up to 3,000 miles (4,828 kilometers). The insects must begin this journey each fall ahead of cold weather, which will kill them if they tarry too long. Monarch butterflies begin life as eggs and hatch as larvae that eat their eggshells and, subsequently, the milkweed plants on which they were placed. (Monarchs are dependent on milkweed plants, which larvae eat nearly exclusively.) Fattening larvae become juicy, colorful caterpillars, then create a hard protective case around themselves as they enter the pupa stage. They emerge as beautifully colored, black-orange-and-white adults. The colorful pattern makes monarchs easy to identify—and that's the idea. The distinctive pattern warns predators that the insects are foul tasting and poisonous. Butterflies that emerge from chrysalides (pupa state) in late summer and early fall are different from those that do so during the longer days and warmer weather of summer. These monarchs are born to fly, and know because of the changing weather that they must prepare for their lengthy journey. Only monarchs born in late summer or early fall make the migration, and they make only one round trip. By the time next year's winter migration begins, several summer generations will have lived and died and it will be last year's migrators' great grandchildren that make the trip. Yet somehow these new generations know the way, and follow the same routes their ancestors took—sometimes even returning to the same tree. Many scientists are concerned about the eastern population of monarchs, which summer east of the Rocky Mountains. This group is occurring in ever smaller numbers, and its survival may be threatened by a series of natural disasters in the Mexican wintering grounds, as well as by reduced acreage of milkweed plants in their summer home. Monarch Butterfly Monarch-Butterfly Monarch-Butterfly Monarch-Butterfly Monarch-Butterfly Monarch-Butterfly Monarch-Butterfly Monarch-Butterfly Monarch Butterfly User Rating: / 10 PoorBest Search Who's Online We have 60 guests online
Take the 2-minute tour × Stack Overflow is a question and answer site for professional and enthusiast programmers. It's 100% free, no registration required. My Javascript and JQuery skills are poor at best and this is ** I have the following three elements in a form : <select name="event_time_start_hours"> <option value="blank" disabled="disabled">Hours</option> <option value="blank" disabled="disabled"> </option> <option value="01">1</option> <option value="02">2</option> <option value="03">3</option> <option value="04">4</option> <option value="05">5</option> <option value="06">6</option> <option value="07">7</option> <option value="08">8</option> <option value="09">9</option> <option value="10">10</option> <option value="11">11</option> <option value="12">12</option> <option value="midnight">Midnight</option> <option value="midday">Midday</option> </select> <select name="event_time_start_minutes"> <option value="blank" disabled="disabled">Minutes</option> <option value="blank" disabled="disabled"> </option> <option value="00">00</option> <option value="15">15</option> <option value="30">30</option> <option value="45">45</option> </select> <select name="event_time_start_ampm"> <option value="blank" disabled="disabled">AM / PM</option> <option value="blank" disabled="disabled"> </option> <option value="am">AM</option> <option value="pm">PM</option> </select> Quite simply, when either 'midnight' or 'midday' is selected in "event_time_start_hours", I want the values of "event_time_start_minutes" and "event_time_start_ampm" to change to "00" and "am" respectively. My VERY poor piece of Javascript says this so far : $(document).ready(function() { $('#event_time_start_hours').change(function() { if($('#event_time_start_hours').val('midnight')) { $('#event_time_start_minutes').val('00'); } }); }); ... and whilst I'm not terribly surprised it doesn't work, I'm at a loss as to what to do next. I want to do this purely for visual reasons for the user as when the form submits I ignore the "minutes" and "am/pm". I'm trying to decide whether it would be best to change the selected values, change the selected values and then disable the element or hide them altogether. However, without any success in getting anything to happen at all I haven't been able to try the different approaches to see what feels right. I've ruled out the obvious things like a duplicate element ID or simply not linking to JQuery. Thank you. share\|improve this question All working now so thank you very much to everyone who answered. I should've realised there were no ID's (I'll blame it on the fact I'm working on someone else's HTML even though there really is no excuse) and thanks for all the JQuery, Javascript tips: I might actually get the hang of it one of these days. – Chris Stevenson Mar 18 '10 at 17:05 5 Answers 5 up vote 1 down vote accepted On your test, you're not comparint the value of #event_time_start_hours to midnight, you're actually setting that value. Try this : $(document).ready(function() { $('#event_time_start_hours').change(function() { if($('#event_time_start_hours').val()=='midnight')) { $('#event_time_start_minutes').val('00'); } }); }); share\|improve this answer Nick's answer is better : selectors use the name instead of elements ID, which you didn't specify in your html, and he gives you the whole reply. – David V. Mar 18 '10 at 17:00 the problem is you're using the id selector (#) but your selects have just a name. share\|improve this answer Your selects are not ID'ed $('#event_time_start_hours') is asking for You have them named so your code should be: $(document).ready(function() { $('select[name=event_time_start_hours]').change(function() { if($('select[name=event_time_start_hours]').val('midnight')) { $('select[name=event_time_start_minutes]').val('am'); } }); }); Oops - fixed 00 to 'am' share\|improve this answer Thank you Ted for being the first to point that one out. I am suitably embarrassed. – Chris Stevenson Mar 18 '10 at 17:02 You need to use a different form of val() to get the current value. And "midday" should be "PM", though you're probably not using that value on the backend anyway. Perhaps it would be better just to hide those dropdowns when the first drop down has "midnight" or "midday" -- in that case, though, the checks could be rolled up into a single if. EDIT: as another user noted, you need to give the selects IDs as well. I'm assuming that you've done that in the code below as it makes the selector shorter. $(document).ready(function() { $('#event_time_start_hours').change(function() { var startHour = $(this).val(); if(startHour == 'midnight') { $('#event_time_start_minutes').val('00'); $('#event_time_start_ampm').val('am'); // or $('#event_time_start_minutes,#event_time_start_ampm').hide(); } else if (startHour == 'midday') { $('#event_time_start_minutes').val('00'); $('#event_time_start_ampm').val('pm'); // or $('#event_time_start_minutes,#event_time_start_ampm').hide(); } }); }); share\|improve this answer Try this: $(function() { $('[name=event_time_start_hours]').change(function() { if($(this).val() === 'midnight' \|\| $(this).val() === 'midday') { $('[name=event_time_start_minutes]').val('00'); $('[name=event_time_start_ampm]').val('am'); } }); }); .val() without parameters returns the value, with a string you're setting the value. share\|improve this answer Your Answer discard By posting your answer, you agree to the privacy policy and terms of service. 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Открывать файлы в режимах “rt” и “wt” \| \| \| \| \| \| \| \| \| \| \| \| Несколько раз здесь, на SO, я видел людей, использующих режимы rt и wt для чтения и записи файлов. Например: с open("input.txt", "rt") в качестве input_file: с open("output.txt", "wt") в качестве output_file: ... Я не вижу задокументированных режимов, но поскольку open( ) не выдает ошибку — похоже, его можно использовать вполне законно. Для чего это нужно и есть ли разница между использованием wt и w и rt против r?
RT Journal Article SR Electronic T1 A synthesis of over 9,000 mass spectrometry experiments reveals the core set of human protein complexes JF bioRxiv FD Cold Spring Harbor Laboratory SP 092361 DO 10.1101/092361 A1 Kevin Drew A1 Chanjae Lee A1 Ryan L. Huizar A1 Fan Tu A1 Blake Borgeson A1 Claire D. McWhite A1 Yun Ma A1 John B. Wallingford A1 Edward M. Marcotte YR 2016 UL http://biorxiv.org/content/early/2016/12/07/092361.abstract AB Macromolecular protein complexes carry out many of the essential functions of cells, and many genetic diseases arise from disrupting the functions of such complexes. Currently there is great interest in defining the complete set of human protein complexes, but recent published maps lack comprehensive coverage. Here, through the synthesis of over 9,000 published mass spectrometry experiments, we present hu.MAP, the most comprehensive and accurate human protein complex map to date, containing >4,600 total complexes, >7,700 proteins and >56,000 unique interactions, including thousands of confident protein interactions not identified by the original publications. hu.MAP accurately recapitulates known complexes withheld from the learning procedure, which was optimized with the aid of a new quantitative metric (k-cliques) for comparing sets of sets. The vast majority of complexes in our map are significantly enriched with literature annotations and the map overall shows improved coverage of many disease-associated proteins, as we describe in detail for ciliopathies. Using hu.MAP, we predicted and experimentally validated candidate ciliopathy disease genes in vivo in a model vertebrate, discovering CCDC138, WDR90, and KIAA1328 to be new cilia basal body/centriolar satellite proteins, and identifying ANKRD55 as a novel member of the intraflagellar transport machinery. By offering significant improvements to the accuracy and coverage of human protein complexes, hu.MAP (http://proteincomplexes.org) serves as a valuable resource for better understanding the core cellular functions of human proteins and helping to determine mechanistic foundations of human disease.
Is your annual physical enough? Here’s what it may miss For many of us, our annual checkup is our only chance to measure our long-term health. Here’s how to get the most out of that visit. Share If you get an annual physical, you may assume you have everything you need to monitor your health. But this once-a-year appointment may be falling short. “Primary care physicians have a set amount of time—usually only about a half hour or less—to cover an incredible amount of ground,” says Ami Kapadia, MD, a functional medicine physician in Portland, Oregon. That’s why it’s ideal to come prepared to get the most out of this annual exam. While most practitioners will ensure you’re up to date on basic tests, such as cancer screenings and bloodwork, there may not be time to look at a holistic picture of your health, she says. “Our current healthcare system is set up to be more reactive than preventative,” Kapadia explains. “It’s set up more to diagnose problems rather than to provide education on prevention.” For instance, with metabolic health, we typically don’t take action until a patient shows signs of metabolic dysfunction that are measurable on conventional lab tests, Kapadia says. “When it comes to Type 2 diabetes, patients often aren’t aware that their blood glucose numbers are less than optimal until they have prediabetes,” she explains. “But the dysfunction may have been there years earlier.” Despite these shortcomings, your annual physical exam is still critical. It’s an insurance-covered opportunity to connect with your doctor and check in on your long-term health. Your physician will check your height and weight, as well as vital signs such as your heart rate and blood pressure. They will also do a head-to-toe exam, checking your mouth and ears, listening to your heart and lungs, and feeling your lymph nodes to look for swelling. Although it’s not a deep dive into any one area, including metabolic health, and physicians will likely be pressed for time, you can often use the appointment to ask questions or even inquire about additional testing. Read on for advice on navigating the standard components of the annual physical to get the most out of the visit, as well as other steps you can take on your own. Are You at Risk for Heart Disease? The Standard Tests The American Heart Association recommends that all adults aged 20 or older have their cholesterol checked at least every four to six years. Most doctors can order a lipid panel to check your cholesterol every year. It typically includes: Total cholesterol. A total cholesterol level of less than 200 mg/dL is the standard for normal. Between 200 to 239 mg/dL is borderline high, and 240 mg/dL or more is high. Levels advisors recommend keeping it below 180 mg/dL. LDL cholesterol. LDL has traditionally been referred to as “bad” cholesterol for its reported role in heart disease risk, though many consider that an oversimplification. Your goal LDL cholesterol is based on your overall risk factors for complications such as a heart attack, including Type 2 diabetes or existing heart disease. If you have no risk factors, standard guidance is that anything under 130 mg/dL is generally acceptable, though Levels advisors recommend keeping it under 100 mg/dL or even 70 mg/dL. HDL cholesterol. Levels below 40 mg/dL raise the risk of heart disease. The ideal is 60 mg/dL or higher. Triglycerides. High triglyceride levels–typically above 150 mg/dL–are associated with an increased risk of cardiovascular disease. Levels advisors recommend aiming for 100 mg/dL or less. These four numbers together can provide a general sense of how your lipid levels may contribute to your heart disease risk, says Kapadia. What Else to Ask About For an even more comprehensive look at your risk of heart disease as it pertains to lipids, we can look at two other numbers: your total cholesterol-to-HDL ratio, and your triglyceride-to-HDL ratio, Kapadia suggests. Both are readily calculated from a basic lipid panel and help look at overall risk of heart disease, she says. Your total cholesterol-to-HDL ratio should be less than 3.5:1 and optimally as low as 2:1. An optimal triglyceride-to-HDL ratio is 2.5:1 or less. You can learn more with our guide to understanding your cholesterol panel, found here. If any of your numbers are outside optimal levels, or you have other risk factors for heart disease, such as a family history of heart disease, being overweight or obese, smoking, or a condition like Type 2 diabetes, Kapadia advises that you may want to ask about the following two additional tests: • ApoB testing measures the amount of a protein, ApoB, prominent in LDL cholesterol. While the optimal levels of ApoB are debated, levels over 110 mg/dL may indicate a higher risk of heart disease. • LDL particle testing measures the number and size of the LDL particles in your blood. A higher amount of small, dense LDL particles may indicate a greater risk of heart disease, even if your total LDL cholesterol is within normal range, says Kapadia. Increased small, dense LDL particles may also be related to insulin resistance. How Is Your Glucose Control? The Standard Tests Your blood glucose levels are an essential indicator of your overall metabolic health, as well as a potential warning sign for metabolic dysfunction. The American Diabetes Association recommends all adults over 35 be screened for Type 2 diabetes at least every three years and more often if they are overweight or have other risk factors, such as a close relative with the condition. Standard testing for diabetes includes fasting glucose and hemoglobin A1c, which provides a snapshot of your glucose control over a three-month period. Levels advisors recommend the following optimal levels: • Fasting glucose: Less than 90 mg/dL • Hemoglobin A1c (HbA1c): Less than 5.5% What Else to Ask About In addition to testing for a fasting glucose level and a hemoglobin A1c, some additional tests can be helpful. “Your A1c and fasting glucose can [fall within the normal range], but you still can be on your way to insulin resistance, which is a precursor to Type 2 diabetes,” Kapadia explains. That’s why she often recommends getting a fasting insulin test. While most practitioners don’t order it routinely, you can request that they add it. Insulin is a hormone that helps move your glucose from your bloodstream into your cells, and it’s vital to keep it at the proper levels to reduce your risk of developing Type 2 diabetes,” she says. An elevated fasting insulin with normal fasting glucose is concerning. There isn’t a scientific consensus on optimal insulin levels, but Levels advisors suggest that fasting insulin stays below 10 microunits per milliliter. Some Levels experts recommend having your medical provider calculate your homeostatic model assessment for insulin resistance (HOMA-IR), indicating how well your body’s insulin response handles glucose. To calculate your HOMA-IR, multiply your fasting glucose (in mg/dL) by your fasting insulin (in uU/mL), and divide it by 405. While the numbers vary by gender, age, and health status, an optimal HOMA-IR is generally under 1. Do You Have Hidden Inflammation? Inflammation can raise your risk of chronic health conditions such as heart disease or certain cancers. One way to help gauge undetected inflammation in your body is to do a simple blood test called a high-sensitivity C-reactive protein (hs-CRP) test. Hs-CRP can be used as one more measurable parameter in helping to determine your risk of heart disease. The Standard Tests Historically, we [doctors] would only order an hs-CRP test if we were monitoring long-term inflammatory conditions, such as rheumatoid arthritis or lupus, for example, or if we were looking for signs of infection, Kapadia says. Over the last two decades, some have also been using hs-CRP as part of the assessment for cardiovascular risk. This measure is most useful for those at intermediate risk of heart disease when we look for more data points to paint an overall picture of their risk. It’s important to note that both acute conditions (like an acute injury, a dental infection, the flu, etc.) and long-term inflammatory conditions can elevate hs-CRP. So, if this value is high in someone otherwise healthy, it’s important to determine if a more acute issue may be the cause and repeat the test periodically. What Else to Ask About Dr. Kapadia also recommends a blood test to measure homocysteine, an amino acid your body uses to make protein, to help diagnose inflammation. High levels of homocysteine are associated with an increased risk of heart disease and can often be treated with B vitamins, which help to break down homocysteine. A normal level is 5–15 mmol/L. Is Your Microbiome Optimal? The gut microbiome is the natural ecosystem of bacteria and other microorganisms that live in your GI tract. They help you digest food, support your immune system, defend against disease-causing bacteria, and may even be protective against conditions such as obesity, Type 2 diabetes, and autoimmune diseases like rheumatoid arthritis. The Standard Tests One test that can help determine the health of your microbiome is a fecal/stool analysis, which sequences the genes of the bacteria and other organisms in your stool. Your doctor can order this type of advanced stool test if you have chronic GI symptoms such as diarrhea or bloating or if they suspect you are having trouble absorbing nutrients. “I use stool testing to look for hidden causes of inflammation and imbalances of microbes such as protozoa or yeast,” says Kapadia. “The test also analyzes your digestive function—for instance, if you secrete enough digestive enzymes for your body to absorb nutrients.” What Else to Ask About In addition, Kapadia suggests breath testing to check for small intestinal bacterial overgrowth (SIBO). In this test, you breathe out after drinking a mixture of glucose or lactulose and water. If there’s a rapid rise in exhaled hydrogen or methane, you may have bacterial overgrowth in your small intestine, she says. Your doctor’s office can do the test, or you can purchase a kit and do it yourself at home. Is Your Nutrient Intake and Absorption Optimal? The Standard Tests “Here are some of the tests Dr. Kapadia finds useful in monitoring nutrient status when getting annual bloodwork: • Ferritin. This is a measure of iron storage in the body. The normal range for blood ferritin is 24 to 336 micrograms per liter for men and 11 to 307 micrograms per liter for women. It’s important to know that iron stores that are both too low and too high can be detrimental to our health. If levels are too high or too low, this requires further investigation to find the cause. • Vitamin B12. Kapadia says many people, particularly older adults, as well as those on certain medications and those with chronic digestive issues, are deficient in this vitamin, which is needed to form red blood cells and the proper nervous system function. Many labs report a range of > 200 pg/mL as normal but give the caveat that some people will experience symptoms at levels less than 400 pg/mL. Kapadia finds it helpful to supplement with B12 when patients test below 400 pg/mL. If your doctor suspects a deficiency, they may also check the levels of a substance in the blood called methylmalonic acid. • Zinc and copper. Zinc helps your body create protein and keeps your immune system running, while copper helps make energy, connective tissue, and blood vessels. Women should have levels of zinc over 70 mcg/dL and copper levels higher than 80 mcg/dL. For men, zinc should be more than 74 mcg/dL, and copper should be higher than 70 mcg/dL. • Vitamin D. At least a quarter of all Americans don’t get enough of this vitamin, which helps your body retain calcium and phosphorus to build bone. The correct dosage depends on your blood levels—people under 70 should aim for 600 IU per day, according to the National Institutes of Health, though Levels advisor Dr. Mark Hyman recommends up to 2000 IU daily. (If you supplement and take more than 2000 IU a day, monitor your blood levels to ensure you’re not taking too much). What Else to Ask About Some practitioners use the NutrEval FMVtest by Genova Diagnostics, which tests a variety of blood and urine biomarkers and assesses the body’s needs for antioxidants, vitamins, minerals, essential fatty acids, and more. A functional medicine physician can order appropriate nutrient lab testing to see where you may be lacking, says Kapadia. Are Your Hormones in Balance? Doctors take different approaches to hormone testing. Levels health advisor Sara Gottfried, MD, for example, thinks it’s important to keep an eye on hormones, including reproductive hormones, even if you’re not actively trying to get pregnant. “If you’re 32 and trying to get pregnant, they’ll check your thyroid, cortisol, testosterone, estradiol, progesterone,” she says. “There is a double standard to be told that it fluctuates too much, and it’s not meaningful when you’re not trying to get pregnant, and yet it is meaningful if you are trying to get pregnant.” Other doctors, like Kapadia, take a different approach. While she orders thyroid hormone testing on most patients, she takes a more targeted approach to further hormone testing. “For instance, if someone is premenopausal and has a lot of menstrual-related symptoms, such as irregular or heavy periods, or if they are perimenopausal and experiencing hot flashes, I’ll order hormone testing routinely,” she explains. For women, hormone evaluations include checking: • Estradiol • Progesterone • Total and free testosterone • FSH and LH • DHEA-S • TSH (thyroid stimulating hormone) For male patients, Kapadia orders testosterone testing if they have symptoms such as low libido, low energy, erectile dysfunction, or trouble building muscle. While testing for cortisol, a stress hormone, can sometimes be helpful, Kapadia doesn’t routinely measure it. “I can usually tell what’s going on with a patient’s cortisol levels based on a thorough clinical history,” she adds. What if your doctor won’t order some of these tests? Some commercial labs allow you to order your own tests directly, no doctor’s visit required. Firms like Everly Well offer home testing for everything from hormone levels to food allergies. (Levels also offers a metabolic blood panel to its members.) Remember that if you self-order tests, insurance is unlikely to cover them, and you may be left with results you don’t understand. Kapadia adds, “I encourage people to go through their doctor for as many tests as possible to get insurance coverage and a proper interpretation of their results,” says Kapadia. “Metabolic health markers, such as glucose and insulin, for example, are usually covered and are inexpensive even when they’re not. And most functional medicine doctors have a cash rate for patients who don’t have good insurance coverage.” Fasting insulin can cost as little as $20, tests like ApoB can be around $60-70, while stool testing and advanced nutrient panels can cost significantly more.
Take the 2-minute tour × Stack Overflow is a question and answer site for professional and enthusiast programmers. It's 100% free, no registration required. I have a model Page, which can have Posts on it. What I want to do is get every Page, plus the most recent Post on that page. If the Page has no Posts, I still want the page. (Sound familiar? This is a LEFT JOIN in SQL). Here is what I currently have: Page.objects.annotate(most_recent_post=Max('post__post_time')) This only gets Pages, but it doesn't get Posts. How can I get the Posts as well? Models: class Page(models.Model): name = models.CharField(max_length=50) created = models.DateTimeField(auto_now_add = True) enabled = models.BooleanField(default = True) class Post(models.Model): user = models.ForeignKey(User) page = models.ForeignKey(Page) post_time = models.DateTimeField(auto_now_add = True) share\|improve this question what is Post relationship to Page? – dm03514 Feb 16 '12 at 1:26 models added for clarity – babonk Feb 16 '12 at 1:37 add comment 3 Answers up vote 2 down vote accepted Depending on the relationship between the two, you should be able to follow the relationships quite easily, and increase performance by using select_related Taking this: class Page(models.Model): ... class Post(models.Model): page = ForeignKey(Page, ...) You can follow the forward relationship (i.e. get all the posts and their associated pages) efficiently using select_related: Post.objects.select_related('page').all() This will result in only one (larger) query where all the page objects are prefetched. In the reverse situation (like you have) where you want to get all pages and their associated posts, select_related won't work. See this,this and this question for more information about what you can do. share\|improve this answer I'm trying to get only the most recent post. page.post_set.all seems to have all of them rather than just that one. how do i get the desired one? – babonk Feb 16 '12 at 6:49 page.post_set.all().latest() in the view or page.post_set.all.latest in the template. You need to specify a get_latest_by field in the Meta class of your model: docs.djangoproject.com/en/dev/ref/models/querysets/#latest – Timmy O'Mahony Feb 16 '12 at 11:02 if you're using that method, what's the point of even getting the most_recent_post=Max('post__post_time')? You could get it from post_set.all.latest.post_time, no? – babonk Feb 16 '12 at 19:48 Yep, that's right – Timmy O'Mahony Feb 16 '12 at 20:12 1 marking this as correct. I ended up doing a raw SQL query. The various solution for django 1.3 and lower aren't very nice. Can't wait for django 1.4 to introduce prefetch_related.. – babonk Feb 16 '12 at 23:41 show 2 more comments Probably your best bet is to use the techniques described in the django docs here: Following Links Backward. After you do: pages = Page.objects.annotate(most_recent_post=Max('post__post_time')) posts = [page.post_set.filter(post_time=page.most_recent_post) for page in pages] And then posts[0] should have the most recent post for pages[0] etc. I don't know if this is the most efficient solution, but this was the solution mentioned in another post about the lack of left joins in django. share\|improve this answer This works, but if there were a way to do it with fewer queries that would be much appreciated. – babonk Feb 16 '12 at 6:54 add comment You can create a database view that will contain all Page columns alongside with with necessary latest Post columns: CREATE VIEW `testapp_pagewithrecentpost` AS SELECT testapp_page., testapp_post. -- I suggest as few post columns as possible here FROM `testapp_page` LEFT JOIN `testapp_page` ON test_page.id = test_post.page_id AND test_post.post_time = ( SELECT MAX(test_post.post_time) FROM test_post WHERE test_page.id = test_post.page_id ); Then you need to create a model with flag managed = False (so that manage.py sync won't break). You can also use inheritance from abstract Model to avoid column duplication: class PageWithRecentPost(models.Model): # Or extend abstract BasePost ? # Page columns goes here # Post columns goes here # We use LEFT JOIN, so all columns from the # 'post' model will need blank=True, null=True class Meta: managed = False # Django will not handle creation/reset automatically By doing that you can do what you initially wanted, so fetch from both tables in just one query: pages_with_recent_post = PageWithRecentPost.objects.filter(...) for page in pages_with_recent_post: print page.name # Page column print page.post_time # Post column However this approach is not drawback free: • It's very DB engine-specific • You'll need to add VIEW creation SQL to your project • If your models are complex it's very likely that you'll need to resolve table column name clashes. • Model based on a database view will very likely be read-only (INSERT/UPDATE will fail). • It adds complexity to your project. Allowing for multiple queries is a definitely simpler solution. • Changes in Page/Post will require re-creating the view. share\|improve this answer add comment Your Answer discard By posting your answer, you agree to the privacy policy and terms of service. Not the answer you're looking for? Browse other questions tagged or ask your own question.
How the 2010 Honda Insight Works By: John Fuller Honda Insight Image Gallery Honda is bringing back the Insight hybrid car, an inexpensive model that still attempts to get drivers good gas mileage. See more pictures of the Honda Insight. Honda Insight Image Gallery Honda is bringing back the Insight hybrid car, an inexpensive model that still attempts to get drivers good gas mileage. See more pictures of the Honda Insight. David McNew/Getty Images When most people bring up the subject of hybrid cars, the Toyota Prius is most likely the first and most prevalent example that comes to mind. It's consistently on year-end best of lists for performance, fuel efficiency and overall satisfaction, and its mid-range price has made it fairly reasonable for most drivers. However, despite the Prius's dominance, the popular car wasn't the first modern hybrid vehicle to hit America's roads and make an impact. That honor belongs to another Japanese carmaker -- Honda Motor Company. In 1999, Honda began producing the 2000 Honda Insight, the first gasoline-electric hybrid to be sold in mass production to U.S. drivers. Advertisement Using a small electric motor to assist the gas engine, the Insight quickly moved to the top of the Environmental Protection Agency's list for best fuel economy. The Insight also introduced Honda's Integrated Motor Assist (IMA), the technology the company would end up using in all of its hybrid powertrains ever since. The system included several aspects that would become the core of hybrid technology, including regenerative braking, nickel-metal hydride battery packs and microprocessor use. Honda stopped producing the Insight in 2006, but the hybrid's hiatus was short-lived. The automakers have refreshed the Insight for the 2010 model year, introducing a new second-generation design to drivers. The hybrid will still use the Integrated Motor Assist system it first brought to the states, but it will also include newer features such as the Ecological Drive Assist System. The system is also known as Eco Assist -- a visual feedback system meant to help drivers improve their fuel economy and overall driving habits. In an effort to entice drivers and rival the Prius, Honda's Insight will be the least expensive hybrid car on the market, priced at $19,800 for the base version. So how does the 2010 Honda Insight work? How fuel-efficient is it? What has Honda changed inside and under the hood for the Insight's second generation? Advertisement 2010 Honda Insight Design The design of the 2010 Honda Insight is similar to that of the Toyota Prius -- but what does it look like under the hood? The design of the 2010 Honda Insight is similar to that of the Toyota Prius -- but what does it look like under the hood? David McNew/Getty Images The Honda Insight's appearance has changed gradually over the years, but compared with its origins the 2010 Insight looks like a completely different car. The original Insight concept began as early as 1997, when Honda introduced the Honda J-VX. A small, lightweight, sporty design, the J-VX was a two-door, four-seater with tapered wheel wells and a flat back. This morphed into the more-recognizable Honda VV, the pre-production prototype of the 2000 Insight. The emphasis was less on sports cars and more on fuel efficiency and aerodynamics, evident in the characteristic extension of the hybrid's aluminum body over the rear wheel wells. Aside from a few refinements in headlamp shape and other physical appearances, the design of the first-generation Insight was basically in place. So how does the new 2010 Honda Insight compare in design to the original? At first glance, a lot of things have changed. Most notably, the rear wheel wells on the 2010 Insight aren't covered up. The grille and headlight design of the new Insight is also significantly different, boasting a more rounded, sectional look than the subtler curves of previous models. Advertisement One thing many reviewers have pointed out while test-driving the 2010 Honda Insight is that is bears a suspicious resemblance to its rival, the Toyota Prius. Both vehicles have a gradually sloping profile, one that begins subtly at the edge of the hybrid's hood, peaks above the front seat headrests, and continues downward in a similar fashion before cutting off quickly at the back. Both cars have four doors (in the case of the Insight, an upgrade from the previous generation's two-door design) and both are hatchbacks, too. There are, however, a few distinctions. The 2010 Insight is considered a compact car, not a mid-sized car like the Toyota Prius, and is therefore considerably smaller. The interiors are generally dissimilar, too, especially in regards to driver controls and feedback. The Insight includes a special Multi-Information-Display (MID) that gives drivers typical statistics like miles per hour and temperature, but also goes the extra mile to provide specific data like fuel economy. The 2010 Insight also comes with a display technology called Eco Assist, designed to give drivers important feedback related to fuel economy. With sophisticated digital visuals, Eco Assist both informs drivers and coaches them along the way. Small details such as sprouting green leaves let you know you're achieving decent fuel economy. Pushing the Econ button near the Insight's dashboard can modify several vehicle settings, allowing drivers to further increase fuel efficiency. Advertisement 2010 Honda Insight Specs The 2010 Honda Insight gets an average of 41 miles per gallon. The 2010 Honda Insight gets an average of 41 miles per gallon. Wes Allison/The Canadian Press/AP Images The Honda Insight has been out of the automotive game for about three years. What's happened during its absence? As Honda brings back its original hybrid car, how does it stack up against other hybrid cars? First, the numbers: The 2010 Honda Insight should be able to achieve 40 miles per gallon (17 kilometers per liter) in the city and 43 miles per gallon (18.3 kilometers per liter) on the highway -- a combined average of 41 miles per gallon (17.4 kilometers per liter), according to the Environmental Protection Agency's estimates. In terms of performance, the Insight features a 1.3-liter, 8-valve, 4-cylinder engine, which delivers 98 horsepower and 123 pound-feet (166.8 newton-meters) of torque. Advertisement Honda has equipped the 2010 Insight with a continuously variable transmission (CVT), a system that exchanges the gears found in typical automatic transmissions for a seemingly simple metal belt that runs between a pair of pulleys. One pulley stays in place, while the other can move with the help of a hydraulic cylinder. When the pulley moves, the belt moves higher or lower on the pulley depending on how fast you're driving, and this changes the gear ratio. It's a lot like changing gears while riding a mountain bike, except in a CVT system, there aren't any steps and there's an infinite number of ratios. This makes a vehicle using a CVT system slightly more efficient, increasing its fuel economy by about one or two miles per gallon (.43 or .85 kilometers per liter). That might not sound like much, but fuel economy is highly dependent on good driving habits -- any extra bit of efficiency engineers can squeeze out of a single system helps, and that's exactly why many automakers like Honda are adding CVTs to their cars. The Insight's engine is, of course, coupled with an electric motor for extra power. Honda's system is known as the Integrated Motor Assist (IMA). The IMA is mounted right between the engine and transmission, and the motor also provides regenerative braking, electric start-up and start-stop technology to improve fuel efficiency during idle stops. Up against similar hybrids like the Toyota Prius, the 2010 Honda Insight isn't quite as efficient -- the Prius gets around 50 miles per gallon (21.3 kilometers per liter), and that's nearly 10 miles per gallon (4.3 kilometers per liter) more than the Insight. Whether or not the Insight's lower price will attract more buyers, however, is yet to be seen. For more information about hybrid cars and other related topics, follow the links on the next page. Advertisement Lots More Information Related HowStuffWorks Articles Sources • ConsumerGuideAutomotive.com. "2000-2006 Honda Insight: overview." Nov. 23, 2008. (Aug. 3, 2009) https://consumerguideauto.howstuffworks.com/2000-to-2006-honda-insight.htm • Healy, James. "2010 Honda Insight hybrid is mostly agreeable." USAToday.com. Jan. 16, 2009. (Aug. 3, 2009) http://www.usatoday.com/money/autos/reviews/healey/2009-01-15-2010-honda-insight_N.htm • Honda.com. "2010 Honda Insight Hybrid - interior features." (Aug. 3, 2009) http://automobiles.honda.com/insight-hybrid/interior.aspx • Honda.com. "2010 Honda Insight Hybrid - performance." (Aug. 3, 2009) http://automobiles.honda.com/insight-hybrid/performance.aspx • Honda.com. "Honda Insight: Integrated Motor Assist." Sept. 15, 2000. (Aug. 10, 2009) http://corporate.honda.com/press/article.aspx?id=2003112040671 • InsightCentral.net. "Insight evolution." 2008. (Aug. 10, 2009) http://www.insightcentral.net/KB/development/index.html • Memmer, Scott. "CVT enters the mainstream." CNET.com. (Aug. 10, 2009) http://www.edmunds.com/ownership/techcenter/articles/45104/article.html • Motor Trend Magazine. "Honda Insight: first drive." March 2009. (Aug. 3, 2009) http://automobiles.honda.com/images/2010/insight-hybrid/downloads/motor_trend.pdf • Russ, Carey. "Driving the 2010 Honda Insight." CNET.com. Jan. 12, 2009. (Aug. 3, 2009) http://reviews.cnet.com/8301-13746_7-10140372-48.html • Siler, Wes. "Report: Honda hustling to refresh Insight." Jalopnik.com. July 22, 2009. (Aug. 10, 2009) http://jalopnik.com/5320320/report-honda-hustling-to-refresh-insight Featured Advertisement Loading...
Caffeine, a widely consumed stimulant found in coffee, tea, chocolate, and medications, has a dual reputation concerning headaches. While it can offer relief for some, it can also act as a trigger for others. Understanding its effects on the body is crucial for managing headache symptoms effectively. • Alertness and Mood Enhancement: Caffeine boosts alertness, concentration, and mood by blocking sleep-promoting neurotransmitters and increasing adrenaline production. • Enhanced Pain Relief: Caffeine accelerates the absorption of painkillers like acetaminophen and aspirin, making them more effective in alleviating headaches. • Vasoconstrictive Properties: By narrowing blood vessels around the brain, it reduces pressure and tension, particularly beneficial for migraines. It’s a common ingredient in headache medications for this reason. • Caffeine Withdrawal Headaches: Regular consumption can lead to dependency, causing severe withdrawal symptoms, including intense headaches, when intake is reduced or missed. • Increased Blood Pressure: Caffeine sensitivity can elevate blood pressure, potentially triggering headaches, particularly for individuals with hypertension. • Sleep Disruptions: Consumption close to bedtime can disrupt sleep patterns, leading to poor sleep quality and increased susceptibility to tension headaches and migraines. • Moderation: Balancing caffeine intake is key, with most people safely consuming 200 to 300 milligrams per day. Sensitivity varies, so monitoring and possibly reducing intake is advisable for those prone to caffeine-related headaches. • Timing: Avoiding it later in the day can improve sleep quality and overall health. Monitoring personal sensitivity helps in adjusting intake time effectively. • Know Your Triggers: Understanding individual responses to caffeine, tracking consumption, and noting associated symptoms help identify triggers and manage intake accordingly. Full Post Starts Next Page >>> Weight Loss Healing Supplements
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METHOD AND SYSTEM FOR CONTROLLING THE QUALITY OF A STAMPED PART A system and method for detecting a defective part and the type of defect formed during stamping operations. The system and method will not only detect the defect but also the nature of the defect and the time at which the defect occurred during stamping operations. Such information is useful not only in quality control but also in isolating a problem which may exist in stamping operations and thus eliminating time for isolating such problems and correcting them. The system and method uses a profile of a properly stamped part to detect a defect, and the root cause of the defect. Skip to: Description · Claims · Patent History · Patent History Description CROSS-REFERENCE TO RELATED APPLICATIONS This application is a divisional of U.S. patent application Ser. No. 13/041,857 filed on Mar. 7, 2011, which is incorporated in its entirety herein by reference. FIELD OF THE INVENTION The invention relates to a system for stamping a part from a blank of material. More particularly, the invention relates to a system and method for detecting abnormalities in the part stamped by a die using forces measured during stamping operations and comparing them with a profile of forces for a properly stamped part. BACKGROUND OF THE INVENTION Stamping operations are done using a die. The die includes an upper die and a lower die, also referenced herein as a slide press and a die cushion respectively. A sheet of material, also referenced in the art as a blank, is placed between the slide and die cushion and the slide and die cushion are pressed against each other so as to form the blank into a desired part. The part may then be visually inspected to ensure that no abnormalities existed during stamping operations. As visual inspection may be time consuming and subject to human error, systems have been put in place to make the inspection automated. For instance, it is known to use the displacement of the press with respect to stamping operations to determine whether or not any flaws occurred during stamping operations. Other methods include measuring compressive and tensile forces on the production part. However, these methods do not take into account the work and force peaks which are transmitted during the stamping operation and thus will not determine the type of the flaw that has occurred. Further, such methods and systems do not provide the type of defect present in the stamped part. Accordingly, it remains desirable to have a system which not only detects whether or not the part is defective but will also provide the type of flaw that occurred thus allowing for operators and management to reconfigure stamping operations to eliminate the flaw. For example, if it is detected that the machine has produced parts which have wrinkles, then the forming characteristics of the forces with respect to time may direct the user as to where in the forming process the wrinkles have been formed. SUMMARY OF THE INVENTION The present invention provides a system and method for detecting a defective part and the type of defect formed during stamping operations. The system and method will not only detect the defect but also the nature of the defect and the time at which the defect occurred during stamping operations. Such information is useful not only in quality control but also in isolating a problem which may exist in stamping operations and thus eliminating time for isolating such problems and correcting them. The method includes the step of establishing a profile. The profile includes characteristics of forces of a properly stamped part. Such characteristics include peak forces applied by the die with respect to a particular point in time during stamping operations. Stamping operations as used herein refers to the process by which die parts are moved together to press a blank and are released from each other so as to free the blank for use in manufacturing. The method further includes the step of measuring the forces of the stamped blank of material and comparing the force characteristics of each of the stamped blanks of material with the profile of the properly stamped part. The profile may further include defective profiles, the defective profile being a profile of an improperly formed part having a particular defect. The defective profiles may include force characteristics for a part formed with a wrinkle, or a split. The existence of the wrinkle or split may be analyzed to determine the root cause of the defect. This root cause may be recorded as part of the defective profile. For instance, a wrinkle may be formed for numerous reasons; however, a particular reason may have a unique force characteristic. Once the root cause is determined, the identified root cause is then associated with that particular wrinkle. The force characteristics of various defects, to include the location of the defect within the part may be recorded and used to for a respective defective profile. Thus, operators overseeing stamping operations may be able to not only identify that the part is defective but the location and nature of the defect. Further, the die operators may be able to easily recognize the root cause of the defect and take corrective measures to minimize manufacturing loss. A system for controlling the quality of a stamped part from a blank of material is also provided. The system includes a die having a slide press and a cushion press. A sensor is mounted to the die and is operable to detect the forces applied to the blank of material. A database having a profile including characteristics of forces of a properly stamped part is used to compare the forces measured during stamping operations. The profile may further include defective profiles. Each of the defective profiles having characteristics of forces which identify the nature of the defect. A processor is in communication with the sensor. The processor compares the forces detected during the stamping of a part and labels the part defective when the detected forces deviate from the profile of the properly stamped part. The processor may further search the database for a defective profile which matches the detected profile so as to identify the nature of the defect. BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a cross-sectional view of a die and a blank; FIG. 2 is a cross-sectional view of FIG. 1 showing the blank being formed; FIG. 3A is an overhead view of the upper die showing sensors formed at each corner; FIG. 3B is a view taken from the bottom of the cushion press showing sensors mounted at each corner of the die cushion; FIG. 4 shows a profile of the forces of a properly formed part for the slide press; FIG. 5 shows a profile of a properly formed part with respect to the die cushion; FIG. 6 is a chart showing the characteristics of forces with respect to the strokes of the die; FIG. 7 is a chart showing forces outside of the force parameters indicating a defect; FIG. 8 is a perspective view of a manufacturing line showing the stamping operations of a part; and FIG. 9 is a diagram showing the steps of a method for detecting a defective part. DETAILED DESCRIPTION OF THE INVENTION Referring to FIGS. 1-8, a system 10 for controlling the quality of a part 12 stamped from a blank of material 14 is provided. The system 10 includes at least one die 16 operable to stamp the blank of material 14 into a desired part 12. A sensor 18 monitors the forces exerted by the die 16. The forces are compared to a profile 20. The profile 20 includes the forces generated during a stamping operation of a properly formed part 12. The profile 20 shows the proper distribution of forces with respect to time. The system 10 labels a part 12 defective when the forces measured differ a predetermined amount from the profile 20. With reference first to FIGS. 1 and 2, an operation of the die 16 is provided. The die 16 has an upper die 22 and a lower die 24, referenced herein as a slide press 22 and a die cushion 24 respectively. The slide press 22 includes tabs 26. The tabs 26 are shown on opposing sides of the slide press 22. The die 16 further includes a pair of binders 28 each having a catch 30. The die cushion 24 is disposed between respective binders 28. The tabs 26 and binders 28 are respectively aligned with each other. The surfaces of the slide press 22 and cushion press 24 are configured to form a predetermined part 12 from a blank. With reference first to FIG. 1, the blank is shown disposed between the slide press 22 and the cushion press 24. The slide press 22 and cushion press 24 are pressed towards each other. The tabs 26 are seated within the catches 30 of the binder 28 and the blank is held in place. The slide press 22 and cushion press 24 are then displaced towards each other so as to stamp the blank of material 14 into a desired part 12. With reference now to FIG. 2, the slide press 22 and cushion press 24 are moved away from each other and the part 12 may then be taken from the die 16. With reference now to FIGS. 3A and 3B, the sensors 18 are shown mounted to respective slide and cushion presses. The sensors 18 are in communication with the processor 32. Preferably the sensors 18 are mounted to the corners of the presses. Such sensors 18 are currently known and used and illustratively include a connection screw. Force is transmitted through the connection screw to the processor 32. A database 34 having a profile 20 of characteristic forces of a properly stamped part 12 may be used to compare forces detected during the stamping of a part 12 to determine the existence of a defect. The profile 20 may be formed through the manufacture of a desired part 12. Specifically, the force characteristics of the part 12 may be collected and compared so as to create the profile 20. Thus the profile 20 may be a historical record of stamped parts 12 which were formed properly. The profile 20 may include force characteristics for both the slide press 22 and the cushion press 24. As used herein, force characteristics relates to the amount of force measured with respect to time, displacement of respective slide and cushion presses with respect to time, and the amount of work done with respect to time. Thus, not only does the profile 20 include the amount of force, but a point along time in which the force was experienced, how much work was done to stamp the part 12, and the whether the slide and cushion presses were in proper position throughout the stamping operation. The profile 20 may further include defective profiles 36. The defective profiles 36 are force characteristics or force characteristics of a particular defect. Thus as parts 12 are stamped, particular defects are recorded. The force characteristics of those defects may be stored in the database 34 as a defective profile 36. For instance, if a part 12 is stamped with a wrinkle or a tear, the force characteristics of the defective part 12 is recorded and stored as a defective profile 36. The processor 32 is in communication with the sensors 18. The processor 32 compares the forces detected during the stamping of the part 12 and compares those forces with the profile 20 of a properly stamped part 12. If the profile 20 matches the detected forces, then the part 12 is identified as being properly formed. If the detected forces do not match the profile 20, then the part 12 is labeled as defective. In cases where a part 12 is labeled defective, the processor 32 further searches the database 34 to determine if the detected forces match any one of the defective profiles 36. Detected forces corresponding to a defective profile 36 is then used to identify the nature of the defect. The processor 32 may further compare other aspects of the stamping operation to the profile 20, to include the work done on the part 12 and the position of the slide and cushion presses with respect to time. The system 10 further includes an encoder 38. The encoder 38 may be disposed on either the slide press 22 or the die cushion 24. The encoder 38 is operable to detect the position of the respective slide or cushion press 22, 24 during stamping operations. [there seems to be more to the use of the encoder 38, in the slides you mentioned how the encoder 38 was necessary, please explain why] With reference now to FIGS. 4 and 5, the profile 20 of a properly stamped part 12 with respect to the slide press 22 and cushion press 24 are provided. The forces are measured with respect to position. As shown, there are peak forces which occur during stamping operations respectively labeled A, B and C. Peak forces indicated by references A and B show the peak forces applied by the slide press 22, whereas peak force labeled C is the peak force applied by the cushion press 24. These force characteristics indicate characteristics which are acceptable for a properly formed part 12. The profiles 20 further include the work done by the respective slide and cushion presses as indicated in the lined portion shown in FIGS. 4 and 5. Thus the profile 20 not only includes identifying when these peak forces are formed with respect to the stamping operations but also how much work each press has done on the part. These five characteristics (the three peak forces and work done by each press) are used to indicate that the part 12 was properly formed. It should be appreciated that these characteristics exemplify a proper forming condition. However, as will be discussed later, other forces and inputs may be used to further narrow what is acceptable as a properly formed part 12. With reference now to FIGS. 6, 7 and 8, a chart showing the forces with respect to the strokes performed by dies 16 placed in succession is provided. FIG. 6 shows a proper profile 20 for the operation of a plurality of dies 16 with respect to the force characteristics and the strokes or presses of the die 16 function. Strokes as used herein refers to the displacement of a slide and cushion press 22, 24 to and from each other. With reference to FIG. 6, a force parameter is provided. The force parameter is indicated by the dashed lines found above and beneath the profile 20. The force parameter is a threshold of acceptable forces during stamping operations. With reference now to FIG. 7, the forces measured during the stamping process of a part 12 is shown. The profile 20 is created by the forces detected by the sensor 18 and transmitted to the processor 32 and processed. In this case it may be seen that there are three instances in which the forces exceed or are outside of the force parameters as defined by the dashed lines. In such a case the system 10 may be shut down. The profile 20 includes peak forces of the slide and cushion presses with respect to time and as stated below, the work performed by both the slide and the cushion presses. However, the profile 20 may also take into account various inputs such as the position of the slide and cushion presses during the stamping operations with respect to time and whether or not oil was placed on the presses or the blank. Other inputs may further include the temperature of the die 16 or the slide and cushion presses or the temperature of the blank for that matter. Thus, a profile 20 of force characteristics for a properly stamped part 12 having a thin film of oil, or without oil may be recorded and used to detect defective parts 12. Thus the profile 20 may be one of many that the user may select based upon the part 12 being stamped, the material used in the blank, and the temperature of the die 16, or whether or not a film of oil was used. The measured forces are compared to the selected profile 20 to determine whether the stamping process produced an abnormal or defective part 12. With reference now to FIG. 8, an operation of the system 10 is provided. The system 10 includes a plurality of dies 16 disposed downstream a steel blank feeder. The steel blank feeder collects blanks and positions them in between the press and cushion slides. Once the blank of material 14 is fed, the slide and cushion presses are displaced towards one another and the tabs 26 of the slide press 22 engage the binders 28 so as to hold the blank in position during forming operations. As the presses are pressed towards one another, the forces exerted by the presses are measured by the sensors 18. The forces are measured throughout the stamping operation of a respective die 16, which is until the presses are displaced from each other. The blank of material 14 is fed downstream the line through each of the dies 16. In the instant case three other die 16 presses are shown downstream the initial die 16. Each die 16 press will have a profile 20 that is characteristic for the work that the die 16 is to do on the blank of material 14. Thus the system 10 may be operable to detect a defective part 12 in any of the die 16 forming processes. Not only is the system 10 capable of detecting a defect, the system 10 may be operable to detect the nature of the defect if a defective profile 36 matches a force characteristic of a formed part 12. The system 10 may further include an automatic shut-off 40 operable to cease operations of the die 16. The automatic shut-off may be actuated when the detected forces deviate from the profile 20 of the properly stamped part 12. With reference to FIG. 8, the automatic shut-off is a button in electrical communication with the system 10. The automatic shut-off 40 may be manually actuated by a press of a button, or may be actuated by the processor depending upon the type or number of defects occurring along the line. An indication as to the nature of the defect may be provided wherein the detected forces match one of the defective profiles 36 stored in the database 34. The existence of the wrinkle or tear may be analyzed to determine the root cause of the defect. This root cause may be recorded as part 12 of the defective profile 36. For instance, a wrinkle may be formed for numerous reasons; however, a particular reason may have a unique force characteristic. Once the root cause is determined, the identified root cause is then associated with that particular wrinkle. Identification of the root cause may reduce manufacturing loss by allowing the operator to go right to the source of the error as opposed to trouble shooting the entire system 10. With reference now to FIG. 9, a method 100 for controlling the quality of a part 12 stamped from the blank of material 14 formed by the operation of a die 16 is provided. The method 100 includes the step of establishing a profile 20 at 102. The profile 20 has characteristics of forces of a properly stamped part 12. The profile 20 may be established by stamping a part 12 and determining if the part 12 meets design specifications and does not include any faults. The characteristic of forces of such a part 12 may be used as the profile 20. The method 100 further includes the step of measuring the forces of a stamped blank of material 14 and comparing the stress characteristics of each of the stamped blank of material 14 with the profile 20 of the properly stamped part 12 at 108. The method 100 proceeds to step 110 where any of the stamped blank material is labeled as being a defective part 12 where the forces deviate from the profile 20 of the properly stamped part 12. The profile 20 may take into consideration the force of the slide and cushion presses and may include establishing a force parameter wherein stamped materials having slide and cushion press 22, 24 forces outside of the side force parameter are labeled as defective. The method 100 may include using other inputs to further define a profile 20. For instance, the method 100 may include the use of the speed of the die 16 operation, the temperature of the die 16, or the existence of a film of oil on the blank of the material to establish the profile 20. Thus, a profile 20 for a properly formed part 12 at 100° Fahrenheit may have different force characteristics than that same part 12 properly formed at 80° Fahrenheit. Likewise, a profile 20 may have different force and force characteristics for the same part 12 where one part 12 is made with a film of oil and the other is not. The method 100 may further include the step of establishing a defective profile 36 at. The defective profile 36 may be established by recording the forces exerted by the slide and cushion press 22, 24 in the formation of a defect in a particular part 12. For instance, the formation of a wrinkle in a part 12 may leave a unique force characteristic. The force characteristics of each of the stamped part 12 may be compared with the defective profiles 36, and the nature of the defect may be provided as shown at step 112. The method 100 may further include the step of recording the root cause and associating the root cause with the defective profile 36 at 106. The root cause of the defect may be analyzed by die 16 operators. Thus the method 100 not only identifies a defect, but also provides the nature of the defect and the root cause. In view of the teaching presented herein, it is to be understood that numerous modifications and variations of the present invention will be readily apparent to those of skill in the art. Likewise, the foregoing is illustrative of specific embodiments of the invention but is not meant to be a limitation upon the practice thereof. It is the following claims, including all equivalents, which define the scope of the invention. Claims 1. A method for detecting a defect in a part stamped from a blank of material formed by the operation of a die, the method comprising: establishing a profile, the profile having a characteristic of forces of a properly stamped part; measuring the forces of the stamped blank of material; comparing the force characteristics of each of the stamped blank of material with the profile of the properly stamped part; and labeling any of the stamped blank of material as being a defective part when the force characteristic deviates from the profile of the properly stamped part. 2. The method as set forth in claim 1, wherein the characteristics of forces includes a slide force and a die cushion force. 3. The method as set forth in claim 2, further including the step of establishing a slide force parameter, wherein stamped material having slide forces outside of the slide force parameter are labeled as defective. 4. The method as set forth in claim 2, further including the step of establishing a cushion force parameter, wherein stamped material having cushion forces outside of the die cushion force parameter are labeled as defective. 5. The method as set forth in claim 1, further including the step of using the speed of the die operation to establish the profile. 6. The method as set forth in claim 1, further including the step of using the temperature of the die to establish the profile. 7. The method as set forth in claim 1, further including the step of placing a film of oil on the blank of material, and establishing the profile based upon the changes in the film of oil placed on the blank of material. 8. The method as set forth in claim 1, further including the step of establishing a defective profile, the defective profile including the force characteristics of a defective part, and the nature of the defect. 9. The method as set forth in claim 8, wherein the defective profile further includes the root cause of the defect. Patent History Publication number: 20150107320 Type: Application Filed: Dec 16, 2014 Publication Date: Apr 23, 2015 Patent Grant number: 9999911 Inventors: Joshua A.H. Walter (Innerkip), Michael J. Gravis (Woodstock), Charles David Apps (Livermore, CA) Application Number: 14/571,845 Classifications Current U.S. Class: Sensing Work Or Product (e.g., By X-ray) (72/17.3) International Classification: B21C 51/00 (20060101); B21D 22/02 (20060101);
Skip to main content Advertisement Modelling cardiac fibrosis using three-dimensional cardiac microtissues derived from human embryonic stem cells Abstract Background Cardiac fibrosis is the most common pathway of many cardiac diseases. To date, there has been no suitable in vitro cardiac fibrosis model that could sufficiently mimic the complex environment of the human heart. Here, a three-dimensional (3D) cardiac sphere platform of contractile cardiac microtissue, composed of human embryonic stem cell (hESC)-derived cardiomyocytes (CMs) and mesenchymal stem cells (MSCs), is presented to better recapitulate the human heart. Results We hypothesized that MSCs would develop an in vitro fibrotic reaction in response to treatment with transforming growth factor-β1 (TGF-β1), a primary inducer of cardiac fibrosis. The addition of MSCs improved sarcomeric organization, electrophysiological properties, and the expression of cardiac-specific genes, suggesting their physiological relevance in the generation of human cardiac microtissue model in vitro. MSCs could also generate fibroblasts within 3D cardiac microtissues and, subsequently, these fibroblasts were transdifferentiated into myofibroblasts by the exogenous addition of TGF-β1. Cardiac microtissues displayed fibrotic features such as the deposition of collagen, the presence of numerous apoptotic CMs and the dissolution of mitochondrial networks. Furthermore, treatment with pro-fibrotic substances demonstrated that this model could reproduce key molecular and cellular fibrotic events. Conclusions This highlights the potential of our 3D cardiac microtissues as a valuable tool for manifesting and evaluating the pro-fibrotic effects of various agents, thereby representing an important step forward towards an in vitro system for the prediction of drug-induced cardiac fibrosis and the study of the pathological changes in human cardiac fibrosis. Background Cardiac fibrosis is a common feature of most myocardial pathologies, including ischaemic cardiomyopathy, inherited cardiomyopathy mutations, metabolic syndrome, diabetes, and ageing [1, 2]. Increased mechanical stress or myocardial injury can trigger cardiac fibrosis, which might contribute to contractile and diastolic dysfunctions and subsequent sudden death [3]. Cardiac fibrosis is characterized by the excess accumulation of extracellular matrix (ECM) components, such as collagen and fibronectin, with the consequent pathological remodelling of ECM, followed by transforming fibroblasts into myofibroblasts [4, 5]. These myofibroblasts have high fibrotic activity marked by the expression of α-smooth muscle actin (α-SMA), resulting in myocardial fibrosis and stiffening, which eventually impairs cardiac function [6, 7]. The exploration of the pathogenesis and therapy development for cardiac fibrosis is hampered by the lack of appropriate experimental models that fully recapitulate human cardiac fibrosis. An improvement of in vitro cell or tissue models may open up new possibilities for disease modelling, drug discovery, and regenerative medicine by providing fast and controllable platforms with high availability and relatively low cost compared to animal models. As yet, few in vitro models of cardiac fibrosis have been developed [8,9,10,11,12]. Most are composed of cardiac cells derived from neonatal mouse and rat hearts and thus have less relevance to human pathophysiology. In addition, most current models of cardiac tissue contain only cardiomyocytes (CMs) and lack other key cell types found in the human heart [13]. Therefore, there is a need for an in vitro human cardiac fibrosis model that possesses the physiologically relevant cell combination and can mimic the three-dimensional (3D) nature of native cardiac tissue. Due to the limited amount of human primary CMs available, CMs derived from human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), have emerged as the most appropriate cell source for modelling the human heart in vitro, with obvious advantages of multiple functionalities and increased throughput [14, 15]. Though hPSC-derived CMs or cardiac progenitor cells represent immature phenotypes in their morphological and other physiological and biochemical properties [16], there is a clear incentive to use hPSC-derived CMs as a foundation to generate 3D human cardiac tissue that can eventually be tailored to patient-specific models of normal and diseased cardiac tissues. However, to date, there are no suitable in vitro cardiac fibrosis model based on hPSC-derived 3D cardiac tissue to study the pathological changes in human cardiac fibrosis and to evaluate novel treatments. In this study, we developed an in vitro model of cardiac fibrosis through hESC-derived 3D cardiac microtissues, composed of CMs and mesenchymal stem cells (MSCs) differentiated from the same hESC line. In the normal adult heart, CMs represent only 30% of the total cell number and the remaining 70% consists of various types of cells, among which cardiac mesenchymal cells are in the majority [17]. Cardiac mesenchymal cells can provide a major precursor population to generate fibroblasts, which mediate scar formation via fibroblast–myofibroblast transition during fibrosis [18,19,20,21,22,23]. Therefore, to engineer a physiologically relevant in vitro cardiac microtissues model, we used MSCs to provide fibroblasts, which can be differentiated towards myofibroblasts after stimulation with pro-fibrotic agents. The molecular and cellular properties of these 3D cardiac microtissues were characterized and the pro-fibrotic consequences were observed after induction with transforming growth factor-β1 (TGF-β1) or other pro-fibrotic mediators. We believe that our disease model using 3D cardiac microtissues may be a suitable in vitro model for studying fibrotic changes in human heart tissue and can potentially contribute to the development of more physiologically relevant preclinical drug discovery platforms. Results Differentiation and characterization of CMs from hESCs To obtain human CMs for the development of a human cardiac microtissue model, we differentiated hESCs, which can provide an unlimited source of starting cells for CM differentiation, into CMs by manipulating Wnt signalling as previously described [24]. At 8 days after the induction of differentiation, the synchronous and spontaneous beating areas were observed (Fig. 1a and Additional file 1: Movie 1). FACS analysis with CM-specific markers (cardiac muscle Troponin T (cTnT)) showed that hESCs were differentiated into CMs with high purity (> 98%) (Fig. 1b). The relative expression of POU5F1, a gene encoding an hESC-specific transcription factor, in differentiated CMs was markedly reduced compared with undifferentiated hESCs (Fig. 1c). By contrast, the expression of genes involved in the formation of sarcomere, the basic unit of myofibrils, including TNNT2, MYH6, and MYH7, and the peptide hormone gene (ANF) secreted in the cardiac muscle were significantly increased (Fig. 1c). We confirmed the expression of cardiac transcription factors (GATA4, MESP1, NKX2.5) in the nucleus (Fig. 1d), and further examined the formation of a myofibril structure responsible for contraction and relaxation of CMs and cardiac sarcomere by immunofluorescence for cTnT, MYL2, MLC2A, and sarcomeric alpha-actinin (SAA) (Fig. 1e). We showed the expression of the gap junctional protein Connexin 43 (Cx43), which plays an important role in the electrical coupling of myocardium [25], in the region of cell–cell interactions (Fig. 1e). Myofibrils stained with MLC2A were connected between the two cells via Cx43 (Fig. 1e). These results showed that CMs differentiated from hESCs were interconnected via Cx43-mediated gap junctions, which have a similar structure to the Cx43 gap junction plaque in the intercalated disc of cardiac muscle [26]. Overall, these data demonstrated that hESC differentiated into CMs with high purity and expressed cardiac function-related proteins, implying that hESC-derived CMs are appropriate cell sources for generation of an in vitro human cardiac tissue model. Fig. 1 figure1 Characterization of CMs differentiated from hESCs. a Representative morphology of undifferentiated hESCs and CMs differentiated from hESCs after 10 days. Scale bars, 100 μm. b Flow cytometry plots of cardiac-specific marker cardiac troponin T (cTnT)+ cells. c Quantitative RT-PCR of hESC-specific marker (POU5F1) and CM-specific markers (TNNT2, TBX5, MYH6, MYH7, and ANF) in undifferentiated hESCs and differentiated CMs. Data are the means±SD of three independent experimental replicates (n = 3). p < 0.01 d Representative immunofluorescent staining of hESC-derived CMs for cardiac-specific transcription factors (GATA4, MESP1, and NKX2.5). Scale bars, 10 μm. e Immunofluorescent staining for sarcomeric proteins (sarcomeric-alpha actinin (SAA), cTnT, MYL2, and MLC2A) and junctional protein (Cx43). Nuclei were stained with DAPI (blue). White arrow indicated Cx43-positive gap junction. Scale bars, 10 μm Generation of 3D human cardiac microtissues In this study, we attempted to simulate the pathological characteristics of cardiac fibrosis by introducing the 3D culture method. When 50,000 hESC-derived single CMs were seeded on round bottom 96-well ultra-low attachment plates, self-aggregation occurred slowly, resulting in uniform-sized cell spheroids (approximately 500 μm in diameter) (Additional file 2: Figure S1A). The created cardiac spheroids were beating at regular intervals and were maintained for more than 2 months (Additional file 3: Movie 2). TGF-β1 signaling has been shown to play important roles in mediating fibrotic responses by regulating ECM remodelling and excessive collagen deposition, which eventually results in cardiac hypertrophy and fibrosis [27]. Therefore, we tested whether cardiac fibrotic phenotypes can be reproduced by treating cardiac spheroids with TGF-β1. However, TGF-β1 treatment for 3 weeks did not show any marked effect on collagen deposition in the cardiac spheroids (Additional file 2: Figure S1B). To establish a microtissue model that ultimately aimed to closely recapitulate human cardiac fibrosis, we used MSCs as a source of fibroblasts, which can be obtained by differentiation from the same hESCs used in differentiating into CMs. The expression of MSC surface markers such as CD105 and STRO-1 was confirmed by immunostaining and over 99% of the differentiated MSCs were in the CD73-positive populations (Fig. 2a, b). Most importantly, we found that most hESC-derived MSCs expressed CD44, a hyaluronic acid receptor and marker of cardiac MSCs [21], as evaluated by immunofluorescence and FACS analysis (Fig. 2a, b). qRT-PCR also confirmed that these differentiated MSCs expressed relevant markers, such as ENG (CD105), NT5E (CD73), and CD44 (Fig. 2c). It has been previously reported that endogenous CD44-positive MSCs contribute to the fibroblast population in myocardial infarction [18]. Fig. 2 figure2 Characterization of MSCs derived from hESCs. a Representative morphology of differentiated MSCs and immunofluorescence staining for MSC-specific markers (CD105, STRO1, and CD44). Nuclei were stained with DAPI (blue). Scale bars, 100 μm. b Histograms of flow cytometry analysis for MSC surface markers (CD73 and CD44). The percentage of CD73+ and CD44+ cells in the total cell population. (c) qRT-PCR analysis of MSC markers (Endoglin (ENG; CD105), Ecto-5-prime-nucleotidase (NT5E; CD73), and CD44) in undifferentiated hESCs and MSCs differentiated from hESCs. Data are the means±SD of three independent experimental replicates (n = 3). p < 0.01 Therefore to develop a reliable 3D cardiac tissue model with increased physiological relevance, hESC-derived CMs were mixed with hESC-derived MSCs similar to those likely to be found in human cardiac tissue. In consideration of a report that cardiac fibroblasts account for 20% of total mass of the myocardium [28, 29] and the considerable variation of the ratio of cell numbers of fibroblasts in heart across studies [30], we tested various ratios of MSCs (5–20%), which are used as a source of fibroblasts in CM-MSC cardiac microtissues. Therefore, a single cell suspension of CMs was mixed at a 5–20% ratio of a single cell suspension of MSCs in round bottom 96-well ultra-low attachment plates to form cardiac microtissues called spheroids. Mixing with MSCs, known ECM-secreting cells, stimulated concentration-dependent formation of self-aggregates into cardiac microtissues (Fig. 3a). The average size of cardiac spheroids comprising hESC-derived CMs and 20% MSCs (597.32 μm ± 7.55) was significantly smaller than that of CM only spheroids (700.00 μm ± 22.65) (Fig. 3b). Cardiac spheroids comprising hESC-derived CMs and 20% MSCs also showed regular contractile activity (Additional file 4: Movie 3). To elucidate the electrophysiological features, we used an MEA system that enables simultaneous, accurate, and real-time recordings from multiple sites in the cell network [31]. MEA analyses revealed that CMs co-cultured with 20% MSCs showed a more regular spontaneous beating pattern with a beating frequency of approximately 49.15 ± 5.66 beats/min at 0 to 1 min than control CMs (17.81 ± 2.80 beats/min), CMs co-cultured with 5% (17.3 ± 0.63 beats/min) or 10% MSCs (26.1 ± 4.24 beats/min) (Fig. 3c). Furthermore, we observed a higher amplitude of field potential (FP) from CMs co-cultured with 20% MSCs compared with control CMs and CMs co-cultured with 5% or 10% MSCs (Fig. 3d). The field potential duration (FPD) of CMs co-cultured with 20% MSCs (616.82 ± 55.91 ms) was significantly increased compared to controls (288.66 ± 19.42 ms), CMs co-cultured with 5% (397.77 ± 20.81 ms) or 10% MSCs (380.23 ± 28.78 ms) (Fig. 3e), suggesting that the addition of 20% MSCs into 3D cardiac microtissues may promote functional improvement. Fig. 3 figure3 Importance of MSCs in generating 3D cardiac microtissue. (a) Representative image of CMs and MSCs differentiated from the same hESC line when co-cultured as 3D spheroids for day 1 and 7. Cardiac spheroid formation was dependent on the ratio of MSCs to CMs. Scale bars, 100 μm. (b) Average diameters of cardiac spheroids. Data are the means±SD of replicates (n ≥ 5). p < 0.01. (c) Representative raw traces of the microelectrode array (MEA) recording from control CMs and CMs co-cultured with 5, 10 and 20% MSCs on day 6 after seeding. (d) The representative traces of averaged field potential (FP) recorded by MEA in the control CMs (n = 14) and CMs co-cultured with 5% (n = 37), 10% (n = 49) and 20% MSCs (n = 48). Open triangle indicates the peak of FP and bilateral arrow indicates the field potential duration (FPD). (e) FPD obtained from the control CMs (n = 14) and CMs co-cultured with 5% (n = 37), 10% (n = 49) and 20% MSCs (n = 48). p < 0.01, p < 0.05 Characterization of 3D human cardiac microtissues A comprehensive assessment of morphological integrity and cellular composition was conducted over cardiac microtissues by using immunofluorescence of key cell type-specific markers and cardiac gene expression analysis. Cardiac spheroids comprising CMs in combination with MSCs (CM-MSC cardiac microtissues) showed a compartmentalized organization in which SAA-positive CMs were concentrated in the center of the spheroid (Fig. 4a), while MSCs that were positive for vimentin, a mesenchymal and pan-fibroblastic marker, were at the periphery of the spheroid (Fig. 4a). Cells in the periphery of the spheroid also expressed the cardiac fibroblast-specific marker DDR2 (Fig. 4b, left panel), a collagen receptor expressed early in the adult heart and in development [32], which was not expressed in hESC-derived MSCs (Fig. 4b, right panel). Furthermore, vimentin and DDR2 double-positive cells emerged, suggesting a transdifferentiation of MSCs into fibroblasts within CM-MSC cardiac microtissues (Fig. 4b, arrows in left panel). Using PCA, we found that global transcriptomic data of CM-MSC cardiac microtissues clustered independently of control cardiac microtissues comprising only CMs, which were isolated from undifferentiated hESCs on principal component 1 (PC1, 51% variance), and became more similar to the transcriptome of the human heart (Fig. 4c). In addition, the transcriptome of the CM-MSC cardiac microtissues was distinct from that of CMs and tended to be closer to the transcriptome of human heart on principal component 2 (PC2, variance 26%) (Fig. 4c). Further analysis of the transcriptomes revealed 746 differentially expressed genes (DEGs) in CM-MSC cardiac microtissues compared with control cardiac microtissues without MSCs. As expected, in the gene-set enrichment with increased genes in CM-MSC cardiac microtissues, a set of genes related to fibroblast function, such as cell signalling, com, and wound healing was predominantly identified (Fig. 4d). Consistently, pathways associated with cardiac muscle function, such as cardiac conduction and membrane depolarization during cardiac muscle cell action potential were also enriched in the DEG analysis (Fig. 4d). To confirm the regulation of genes related to cardiac muscle function by mesenchyme, the expression of the related genes was confirmed by qRT-PCR. In agreement with the microarray results, there was increased expression of genes related to cardiac function such as SGCD, which plays a role in dystrophin complex stabilization, MYL1, encoding the myosin light chain involved in cardiac conduction, and SCN7A, SCN1B, KSNJ2, and KCNE4, encoding voltage-gated ion channel related proteins (Fig. 4e). These results suggest that CM-MSC cardiac microtissues not only acquired fibroblastic functions by changing cellular composition but might also improve cardiac function through myocyte-mesenchyme interactions. Fig. 4 figure4 Cellular and molecular assessment of 3D cardiac microtissue. a Immunofluorescent staining for cardiac-specific marker (sarcomeric-alpha actinin; SAA) and MSC/pan-fibroblast-specific marker (vimentin; VIM) to visualize cell distribution in CM-MSC cardiac microtissue on day 14. Nuclei were stained with DAPI (blue). Scale bars, 50 μm. b Cardiac microtissue sections were co-immunostained with anti-vimentin (green) for MSC/pan-fibroblast-specific marker and anti-DDR2 (red) for the cardiac fibroblast marker (left panel). Cells positive for both vimentin/DDR2 occurred within cardiac microtissue (arrows), suggesting a transdifferentiation of MSCs into fibroblasts. hESC-derived MSCs were analyzed by immunofluorescence using anti-vimentin (green) and anti-DDR2 (red) (right panel). c Principal component analysis (PCA) based on total gene expression in undifferentiated hESC (n = 3), hESC derived CMs (hESC-CM) (n = 2), CM-MSC cardiac microtissue (hESC-CM + hESC-MSC) (n = 3), and human heart (n = 1). d Reactome pathway terms enriched in up-regulated DEGs (> 2-fold change) in the transcriptome of CM-MSC cardiac microtissues, compared to that of hESC-CM. e qRT-PCR analysis of cardiac function-related genes, including SGCD, MYL1, SCN7A, SCN1B, KCNJ2, and KCNE4. Data are the means±SD of three independent experimental replicates (n = 3).p < 0.01 TGF-β1-induced fibrosis in CM-MSC cardiac microtissues To develop a fibrosis model using CM-MSC microtissues, they were treated with the pro-fibrotic growth factor TGF-β1 for 2 weeks. Treatment of TGF-β1 induced irregular beating patterns in cardiac microtissues (Fig. 5a and Additional file 5: Movie 4). To observe collagen deposition, Masson’s Trichrome staining was performed in sections of TGF-β1-treated CM-MSC microtissues in which a thicker collagen layer was detected after increasing the days of TGF-β1 treatment as seen in fibrotic disease tissues (Fig. 5b). A highly consistent pattern of results was obtained by conducting repetitive experiments (Additional file 6: Figure S2). CMs of CM-MSC cardiac microtissues treated with TGF-β1 exhibited significantly increased apoptosis as evidenced by increasing SAA/cleaved caspase-3 double-positive cells (Fig. 5c, d). The average sphere size was found to be approximately 10% smaller in TGF-β1-treated CM-MSC microtissues (554.17 μm ± 14.87) compared with the control (610.36 μm ± 8.06) (Fig. 5e), which is the result of CM apoptosis. Furthermore, TGF-β1 treatment also promoted the transdifferentiation of cardiac fibroblasts into myofibroblasts, a major cause of abnormal collagen secretion, as shown by the increased number of α-SMA-positive cells (Fig. 5f). In the TGF-β1-treated group, α-SMA-positive cells increased over time (Fig. 5f). Most importantly, cells that were positive for both vimentin and α-SMA were observed in the TGF-β1-treated group, strongly indicating that cells of fibroblast origin had transdifferentiated into myofibroblasts (Fig. 5g, arrows). These results demonstrated that treatment of TGF-β1 in CM-MSC cardiac microtissues can recapitulate collagen deposition, enhanced apoptosis in CMs, and myofibroblast differentiation, which are typical pathological phenotypes observed during in vivo myocardial fibrosis. Fig. 5 figure5 Recapitulation of phenotypes observed in TGF-β1-induced cardiac fibrosis using 3D cardiac microtissue. a Representative images of CM-MSC cardiac microtissue with or without 5 ng/ml TGF-β1 treatment for 2 weeks (left panel). Representative trace obtained by plotting z-axis profile of cardiac microtissue beating (right panel). b Masson’s Trichrome staining of CM-MSC cardiac microtissue sections at serial time points after TGF-β1 treatment. Note the extensive interstitial fibrosis represented by the blue stains. Scale bars, 100 μm. c Immunofluorescent staining of apoptotic CMs in cardiac microtissue with an apoptosis-specific marker (Cleaved caspase 3; Cl-Casp3) and cardiac specific marker (sarcomeric-alpha actinin; SAA). White arrow indicates cells co-stained with SAA and Cl-Casp3. d Percentage of apoptotic CMs by quantifying ratio of Cl-Casp3 positive cells per number of DAPI-stained cells. Data are the means±SD of replicates (n = 3). p < 0.01. e Average diameters of cardiac spheroids. Data are the means±SD of replicates (n ≥ 11). p < 0.01. f Immunoperoxidase staining of myofibroblast-specific marker (alpha-smooth muscle actin; α-SMA) in cardiac microtissues after TGF-β1 treatment. g CM-MSC cardiac microtissue sections were co-immunostained with anti-vimentin (green) and anti-α-SMA (red). Cells undergoing fibroblast-to-myofibroblast transdifferentiation were positive for both vimentin and α-SMA (arrows). Nuclei were stained with DAPI (blue). Scale bars, 50 μm Furthermore, we analysed global transcription levels to investigate the molecular phenotype of TGF-β1-induced fibrosis in CM-MSC cardiac microtissues. Gene set enrichment analysis (GSEA) by biological process, hallmark and cellular localization was performed to identify functional biological pathways in TGF-β1-induced fibrosis models (Fig. 6 and Additional file 7: Figure S3). Consistent with the cellular phenotype shown in Fig. 5b, the extracellular structure organization was ranked as the upregulation pathway in the biological process analysis (Fig. 6a). Likewise, hallmark analysis showed an increase in gene sets related to epithelial mesenchymal transition (EMT) and TGF-β signalling (Fig. 6a). In agreement with these data, GSEA by the cellular location demonstrated that proteinaceous extracellular matrix or the genes located on the basement membrane were enriched in the TGF-β1-induced fibrosis model (Additional file 7: Figure S3A). These analyses suggested that ECM deformation by TGF-β signalling was a major pathway in the fibrosis model. On the other hand, gene sets related to the respiratory chain or inner mitochondrial membrane proteins were enriched in the control cardiac microtissue model (Additional file 7: Figure S3B). Consistent with these data, GSEA by the hallmark revealed that oxidative phosphorylation and fatty acid (FA) metabolism were decreased in the fibrosis model (Fig. 6b). These results raised the possibility that not only the accumulation of ECM as shown in Fig. 5b but also mitochondrial dysfunction may be induced in the TGF-β1-induced cardiac fibrosis model. To assess this possibility, we performed immunofluorescence of TOM20 to visualize the mitochondrial structure, which is closely related to mitochondrial function. Consistent with previous studies [33, 34], the fragmented mitochondrial morphology was observed in cardiac microtissue treated with TGF-β1, suggesting that mitochondrial dysfunction was induced in the cardiac fibrosis models (Fig. 6c). In addition, as FA metabolism is a predominant metabolic pathway in normal heart [35], changes in the expression of FA metabolism-related genes in fibrosis models may adversely affect the normal function of myocardial cells. Fig. 6 figure6 Molecular signatures of TGF-β1-induced cardiac fibrosis models using 3D cardiac microtissue. a List of gene set enrichment subsets in TGF-β1 induced cardiac fibrosis model. Enrichment plot of top ranked subset signature by BP (biological process) and hallmarks; extracellular structure organization (BP) and epithelial mesenchymal transition (EMT) (hallmarks) b List of gene set enrichment subsets in control cardiac microtissue sample. Enrichment plot of top ranked subset; oxidative phosphorylation and fatty acid metabolism. Normalized enrichment scores (NESs) were calculated to account for differences in gene set size and correlations between gene sets and the expression dataset. The false discovery rate (FDR) q-value represents the probability of false discovery associations for a given NES. FDR q-value lower than 0.25 was considered significant. c Immunofluorescence staining of mitochondrial-specific marker (TOM20). Nuclei were stained with DAPI (blue). Scale bars, 10 μm Drug-induced fibrosis in CM-MSC cardiac microtissues To further explore the applicability of CM-MSC cardiac microtissues to drug-induced cardiac fibrosis models, we examined reference pro-fibrotic mediators, including bisphenol A, aldosterone and metoprolol, which are known to induce cardiac fibrosis in vivo [36,37,38]. To explore drug responsiveness in the CM-MSC cardiac microtissue model, tissues were treated with each pro-fibrotic mediator at a concentration of 10 μM for 2 weeks. qRT-PCR showed an increase in TGF-β1-responsive genes, including SERPINE1, CSNK2A, CSNK2B, and CTGF, and collagen genes, including COL1A1, COL1A2, and COL3A1, although variations of responsiveness to each drug were observed (Fig. 7a). An increase in collagen deposition was seen in each compound-treated group compared with control CM-MSC cardiac microtissues (Fig. 7b). Hence, the contribution of pro-fibrotic mediators in transdifferentiation into myofibroblasts was examined in CM-MSC cardiac microtissues, as shown by immunohistochemistry with anti-α-SMA antibody (Fig. 7c). Among the three reference compounds, aldosterone was found to be the most effective for inducing fibrosis in our CM-MSC cardiac microtissues. The size of the spheres was significantly reduced after treatment with each compound (Fig. 7d). CMs of CM-MSC cardiac microtissues treated with reference compounds also exhibited significantly increased apoptosis as evidenced by increasing SAA/cleaved caspase-3 double positive cells (Fig. 7e, f). Moreover, mitochondrial phenotypes were confirmed in the drug-treated cardiac microtissues through TOM20 immunohistochemistry, indicating that mitochondrial fragmentation was increased by treatment with each drug (Fig. 7g). We examined whether treatment with pro-fibrotic drugs could directly influence CMs and the mitochondrial structure. There was no apparent apoptotic response or mitochondrial fragmentation in CMs (Additional file 8: Figure S4). Overall, 3D CM-MSC cardiac microtissues were able to recapitulate the pathological phenotypes of cardiac fibrosis, with differences in fibrotic responses depending on the drug. Therefore, the fibrosis model using CM-MSC cardiac microtissues will be an applicable model system for investigating mechanistic insights into fibrotic diseases and in vitro screening of compounds for drug-induced cardiac fibrosis. Fig. 7 figure7 3D cardiac microtissue for in vitro assessment of drug-induced cardiac fibrosis. a qRT-PCR analysis for mRNA expression for fibrosis-related collagen genes (Col1a1, Col1a2, and Col3a1) and TGF-β responsive genes (SERPINE1, CSNK2A, CSNK2, and CTGF) in CM-MSC cardiac microtissues independently treated with 10 μM concentration of each pro-fibrotic drug for 14 days. Data are the means±SD of three independent experimental replicates (n = 3). p < 0.01, p < 0.05. b Masson’s Trichrome staining to detect collagen deposition in CM-MSC cardiac microtissues following treatment with pro-fibrotic drugs for 14 days. Scale bars, 100 μm. c Immunoperoxidase staining of myofibroblast-specific marker (alpha-smooth muscle actin; α-SMA) in cardiac microtissues after treatment of pro-fibrotic drugs. Scale bars, 50 μm. d Average diameters of cardiac spheroids. Data are the means±SD of replicates (n ≥ 6). p < 0.01. e Immunofluorescent staining of apoptotic CMs in cardiac microtissue with an apoptosis-specific marker (Cleaved caspase 3; Cl-Casp3) and cardiac-specific marker (sarcomeric-alpha actinin; SAA). White arrow indicates cells co-stained with SAA and Cl-Casp3. Scale bars, 50 μm. f Percentage of apoptotic CMs by quantifying ratio of Cl-Casp3 positive cells per number of DAPI-stained cells. Data are the means±SD of replicates (n = 4). p < 0.01. g Immunofluorescence staining of mitochondrial-specific marker (TOM20). Nuclei were stained with DAPI (blue). Scale bars, 10 μm Discussion The goal of engineering a 3D cardiac tissue model is to provide new opportunities for in vitro cardiac modelling with physiologically relevant cell types and microenvironment. The human 3D cardiac microtissue described in this study is expected to improve our knowledge about clinically relevant information regarding cardiac fibrosis and disclose advanced features of cardiac fibrosis. It is now widely believed that cultures of single cell types are very simplistic models, because they do not mimic the complexity and heterogeneity of human tissues. The normal heart is composed of several different cell types, such as CMs, endothelial cells, smooth muscle cells and fibroblasts, which play a role in the development of the heart and its normal functions [39]. Even within the in vitro environment, non-CMs influence the improvement of CM phenotypes and tissue architecture only in 3D cultures [40, 41]. These results are consistent with observations that most cell types do not exhibit the full spectrum of tissue-specific functionality in 2D cultures, since they do not maintain their phenotypes and patterns of gene expression precisely in the different environments compared with native tissue [42]. Therefore, to mimic the native cardiac tissue, our in vitro 3D co-culture strategy that includes relevant cell types appears most appropriate. In this study, we have established for the first time a human 3D cardiac microtissue containing hESC-derived CMs in combination with hESC-derived MSCs. During heart development, fibroblasts, which are important cells that produce fibrotic ECM proteins [43], arise from multipotent progenitor cells, especially MSCs [19,20,21, 23, 44]. The cellular origins of cardiac fibroblasts are varied, including epithelial-mesenchymal transition (EMT) of epicardium cells, endothelial-mesenchymal transition (endo-MT) of epithelial cells, and MSCs [45]. Fibroblasts are connective tissue cells of mesenchymal origin that synthesize and secrete the main components of ECM, such as interstitial collagen and fibronectin [46, 47]. Fibroblasts also play an important role in fibrotic tissue formation by conversion into myofibroblasts under pathological conditions [48]. However, there is currently no standardized protocol for the reliable differentiation of hPSCs into cardiac fibroblasts, whereas, the differentiation methods of hPSCs into MSCs are well established [49, 50]. It has been also reported that MSCs can transdifferentiate into fibroblasts in vitro and in vivo [51]. Cardiac MSCs play an important role in preserving normal cardiac functions, as well as in cardiac remodelling at various pathological stages, by responding early to myocardial infarction [52]. Therefore we developed 3D cardiac microtissues consisting of CMs in combination with MSCs as a source of fibroblasts for developing the in vitro cardiac fibrosis model. The generation of cardiac spheroids with hESC-derived CMs and MSCs has an advantage in terms of the spheroid formation time. As shown in Fig. 3a, aggregates started to form only 1 day after seeding CMs mixed with 20% MSCs, and compact and circular spheroids were successfully generated, whereas loose aggregates with poor cell-cell contacts were formed when CM was cultured only in 3D. Consistent with our findings, Ong et al. have shown that multicellular cardiac spheres are generated from hiPSC-CMs only when they are mixed with human adult ventricular cardiac fibroblasts and human umbilical vein endothelial cells [53]. Furthermore, increased expression of sets of genes associated with ECM organization and cell adhesion (Fig. 4d), which are known to be essential for sphere formation [54], in CM-MSC cardiac spherical microtissues allows more rapid sphere formation in comparison to CM spheroids. DDR2 is known to be selectively expressed in cardiac fibroblasts [45, 46, 55], although it appears to be also expressed in pericytes/vascular smooth muscle cells [56]. As shown in Fig. 4b, all of vimentin-positive MSCs were not co-expressed with DDR2, whereas some cells within CM-MSC cardiac spheroids were double-positive for vimentin and DDR2, suggesting a transdifferentiation of MSCs into fibroblasts within CM-MSC cardiac spheroids. Our findings are supported by a report that CD44-positive MSCs served as a major precursor pool for fibroblasts that mediate scar formation and wound healing after myocardial infarction [18]. In addition, bone marrow-derived MSCs [57, 58] and perivascular-resident MSCs [59, 60], as well as cardiac fibroblasts, which are known to be direct sources of myofibroblasts and MSCs, also exhibit myofibroblastic phenotypes in vitro in response to TGFβ treatment [61]. As in previous cardiac fibrosis models, mouse and rat neonatal CMs are commonly used for developing engineered cardiac tissue because terminally differentiated adult CMs cannot proliferate sufficiently in vitro [8, 10, 12]. In this study, CMs derived from hESCs were well characterized by the expression of cardiac function-related proteins (Fig. 1a-e). Particularly, CMs were connected to each other by the intercalated disc structure by using immunofluorescence staining of Cx43, the most abundant cardiac gap junction protein (Fig. 1e). Cx43 expression was observed as dots in the intercellular connected regions, in which the ends of myofibrils were connected, consistent with a previous study [62]. hPSC-derived CMs have been widely used to perform drug and toxicity testing in vitro [63, 64]. Furthermore, CMs differentiated from hiPSCs generated from patients with genetic cardiac diseases also allow for the generation of CMs carrying the genetic background of a given patient, creating highly tailored models for cardiac diseases in vitro [65]. Recently, a cardiac model of multicellular systems with several cell types, including CMs, endothelial cells, and fibroblasts, was developed to engineer a physiologically relevant cardiac tissue model using a 3D culture system, various biomaterials and bioprinting technology [10, 13, 53, 66,67,68,69]. These multicellular spheroids and 3D models will be applied for developing cardiac fibrosis models by introducing external fibrotic signals. Moreover, when developing in vitro cardiac fibrosis models for personalized drug efficacy and toxicity testing, the iPSC technology applied in this study enables the creation of in vitro disease models in which various cell types with the same genetic background can be assembled. Therefore, disease models using patient-specific cells can be used as platforms to explore new therapies by providing individually optimized treatments. In this work, we established a 3D in vitro model of cardiac fibrosis using 3D cardiac microtissues by treatment with TGF-β1, which is a potent stimulator that promotes the differentiation and proliferation of myofibroblasts during the course of this disease [70]. Our in vitro model, long-term treatment of TGF-β1 induces irregular beating patterns (Fig. 5a), an increase in the accumulation of fibrillar collagen deposition (Fig. 5b), elevated expression of α-SMA, conversion to myofibroblasts (Fig. 5f), increase in the rate of apoptosis of CMs (Fig. 5c, d), and disruption of the mitochondrial network as shown by transformation of filamentous into punctate mitochondria (Fig. 6c), consistent with the pathological changes during cardiac fibrosis [70, 71]. However, we did not detect increased proliferation of MSC or transdifferentiated myofibroblasts in response to TGF-β1 treatment (data not shown), which may have been due to the inclusion of serum-free medium in the cardiac sphere culture conditions. Moreover, we examined whether 3D cardiac microtissues could be used to detect and monitor cardiac fibrosis using known pro-fibrotic mediators. After a 2-week exposure to pro-fibrotic mediators, including bisphenol A, aldosterone and metoprolol, 3D cardiac microtissues display fibrotic features (Fig. 7a-g), such as an up-regulation of pro-fibrotic and TGF-β1-responsive genes, collagen secretion and deposition; increased α-SMA expression; and disorganization of the mitochondrial network, leading to mitochondrial fragmentation, without significant differences compared to TGF-β1-treated cardiac microtissues. These results suggested that our 3D cardiac model can be applied to identify thus far unknown fibrotic compounds and to provide a platform for studying mechanisms of cardiac fibrosis. However, the expression levels of TGF-β1 responsive genes varied among pro-fibrotic drugs (Fig. 7a), suggesting that there may be different detailed molecular mechanisms for the activation of the TGF-β1 pathway. The link between pro-fibrotic drugs and the TGF- β1 pathway is very complex since different molecular mechanisms are involved. Therefore, this research model could provide the basics for studying the development of cardiac fibrosis in response to the different drugs. However, we recognize the limitations of this in vitro model in recapitulating human diseases, as it lacks adequate blood flow and other physiological factors, such as endothelial and inflammatory cells, which are found in the native heart. There is also a need to apply natural and synthetic biomaterials mimicking native ECMs, as biomaterials play a major role in constructing 3D tissue models via supporting cell attachment and alignment and providing tissue-relevant stiffness [72]. Therefore, the further introduction of various cell types present in the heart, application of microfluidics, and use of tissue engineering strategies can be exploited in the development of physiologically relevant in vitro human heart tissue. Conclusions In this study, we report a simplified cardiac microtissue comprising CMs and MSCs derived from hESCs to study cardiac fibrosis in a 3D spheroid platform. Our study highlights the importance of increasing the cellular complexity by adding the appropriate amount of CD44-positive MSCs which can contribute to cardiac fibrosis by generating fibroblasts to better model human cardiac tissues in vitro. Because our cardiac tissue model can be adapted to mimic various aspects of cardiac fibrosis, it cannot only be used to provide further insights into the mechanisms underlying cardiac fibrosis but can also potentially contribute to the development of in vitro assay systems for testing pro-fibrotic compounds and new anti-fibrotic therapies. Methods Cell culture H9 hESCs were obtained from the WiCell Research Institute (Madison, WI, USA) and maintained as described previously [73]. This study using human embryonic stem cell lines was approved by the Public Institutional Bioethics Committee designated by the Ministry of Health and Welfare (MoHW) (Seoul, Republic of Korea; IRB no. P01–201409-ES-01). Differentiation of hESCs into human CMs Human cardiomyocytes were differentiated as described previously [24]. hESCs were transferred to Matrigel (BD Bioscience, San Jose, CA)-coated plates with mTESR medium (Stem Cell Technologies, Vancouver, Canada) without feeder cells; then, cells were grown to 90% confluency. For mesoderm induction, hESCs were treated with 6 μM CHIR99021 (Tocris, Bristol, UK) for 2 days in Cardiac Differentiation Medium (CDM) consisting of RPMI1640 supplemented with 50 μg/ml L-ascorbic acids (Sigma-Aldrich, St. Louis, MO, USA) and 500 μg/ml recombinant albumin (Sigma-Aldrich). Cardiac specification was performed by treating with 2 μM of Wnt-C59 (Selleck Chemicals, Houston, TX, USA) for another 2 days in CDMs. Subsequently, the cells were replaced with CDM medium once every two days until beating CMs were observed. Differentiated CMs were maintained in RPMI-B27 medium consisting of RPMI1640 supplemented with 50 μg/ml L-ascorbic acids and 1XB27 (Thermo Fisher Scientific, Waltham, MA, USA). Differentiation of hESC into human MSCs Human MSCs were differentiated from hESCs as described previously [74]. hESCs were transferred to Matrigel-coated plates with mTESR medium (Stem Cell Technologies) without feeder cells; then, cells were grown to 50% confluency. For mesenchymal differentiation, medium was changed to MEM-alpha supplemented with 10% FBS and 5 ng/ml bFGF (R&D Systems, Minneapolis, MN, USA) once every two days. Cells were transferred to a gelatine-coated dish every week. Generation of CM-MSC cardiac microtissues The differentiated CM and MSC cells were dissociated into single cells by 0.25% trypsin treatment, and the number of cells was counted with a haemocytometer. The CMs and MSCs were mixed at a ratio of 4:1, and 50,000 cells per well were seeded in a 96 well ultra-low attachment plate with RPMI-B27 media supplemented with 10% FBS. After the sphere formation was completed within 2–3 days, the medium was replaced with RPMI-B27 and the medium was replaced every 2–3 days. To develop the in vitro fibrosis model, 5 ng/ml TGF-β1 or 10 μM concentration of each cardiotoxic chemical, including Bisphenol A, Aldosterone, and Metoprolol (all reagents from Sigma-Aldrich) were treated independently with the indicated concentration of drugs as described in previous reports [36,37,38] and the samples were harvested for analysis after 2 weeks. FACS analysis To verify the CM and MSC differentiation efficiency, FACS analysis was performed as described previously [75]. The differentiated CMs and MSCs were treated with 0.25% trypsin, dissociated into single cells, and incubated in dPBS containing 2 mM EDTA and 2% FBS for 10 min. After that, antibodies to the surface markers of each cell (Additional file 9: Table S1) were diluted 1:50 and reacted for 30 min. After washing twice with FACS buffer, FACS analysis was performed using Accuri C6 flow cytometry (BD Biosciences) and analysed using FlowJo V10 software (TreeStar, USA). Processing of cardiac microtissue, immunostaining and MT staining Cardiac microtissues were fixed by 4% paraformaldehyde (PFA), cryo-protected in 10%~ 30% sucrose solution and frizzed after embedding in optimal-cutting-temperature (OCT) compound (Sakura Finetek, Tokyo, Japan) as described previously [76]. Frozen section were prepared by cutting at a thickness of 10–15 μm using a cryostat microtome. The frozen sections were permeablized with 0.1% Triton X-100 for immune-staining, blocked with 3% BSA, and incubated with the primary antibody (Additional file 9: Table S1). For immunofluorescence detection, cells were incubated with a fluorescence-conjugated secondary antibody. For immunoperoxidase detection, the sections were incubated with the biotinylated secondary antibody and then followed using the VECTASTAIN elite ABC kit (Cat. NO. PK-6100, vector laboratories) and DAB substrate kit (Cat. NO. SK-4100, Vector Laboratories, Burlingame, CA, USA). Staining of collagen fibres was performed using Masson’s Trichrome staining kit (Cat no. 25088–1, Polysciences, Inc. Warrington, USA). Quantitative RT-PCR (qPCR) To compare mRNA expression levels, total RNA was extracted according to the manufacturer’s protocol using the easyBLUE RNA extraction kit (iNtRON Biotechnology, Inc., Republic of Korea). cDNA was synthesized using the SuperScript™ IV First-strand Synthesis System (Cat.NO.1891050, Thermo Fisher Scientific), and the cDNA was used as a template for real-time qPCR. PCR reactions were performed three times independently using Fast SYBR ™ Green Master Mix (4,385,612, Applied Biosystems, Foster City, CA, USA). The sequence of the gene-specific primers are presented in the Additional file 10: Table S2. Transcriptome analysis by microarray For global transcriptome analysis, cDNA microarray was performed using an Agilent Human GE (V2) 4 X 44 K chip as described previously [77]. Data were normalized by using GeneSpring software, and differentially expressed genes (DEGs) were analysed by MultiExperiment Viewer software (MeV version 4.9.0; http://mev.tm4.org/). Pathway enrichment analysis was performed by Reactome FI plugin of the Cytoscape software (Version 3.2.0, www.cytoscape.org), and gene set enrichment analysis was performed by GSEA 2.2.4 (http://software.broadinstitute.org/gsea/index.jsp). The H (hallmark genesets) subsets of MSigDB (50 gene sets), C5_BP (GO biological process, 4436 gene sets) and C5_CC (GO cellular component, 580 gene sets) were used in this study [78]. Beating rate analysis of CM-MSC microtissue by tracing of microscopic video files Beating rates of CM-MSC microtissue were recorded on a microscope (IX83, Olympus, Japan) with an on-stage incubator (Live Cell Instrument, Seoul, South Korea) at 37 °C and 5% CO2 to maintain the physiological condition. The plot z-axis profile of each region of interest (ROI) was calculated by Image J software (https://imagej.nih.gov/ij/) program [79]. Field potential recordings using a multi-electrode array (MEA) system The hESC-derived CMs were plated on a Matrigel-coated 12-well MEA plate (4 × 104 cell per well) (AXION, Atlanta, GA, USA) in RPMI-B27 media supplemented with 5% FBS. At 24 h after seeding, the medium was replaced with RPMI-B27. Field potentials were recorded at day 6 after seeding using Axion BioSystems’ Maestro MEA systems set as 37 °C and perfused with 5% CO2, 20% O2, and 75% N2. Field potential signals were recoded with Axion BioSystems’ Integrated Studio (AxIS) software version 2.3 and analysed with the Axion Cardiac Data Plotting Tool. Statistical analysis and graph drawing Whether treatments produce significantly different results was evaluated by statistical analysis. The unpaired t-test (Figs. 1c, 2c, 3b, e, 4e, 5d, and e) and one-way ANOVA followed by Dunnett’s multiple comparisons test (Fig. 7a, d, and e) were performed using GraphPad Prism version 6.00 for Windows (GraphPad Software, Inc., USA). Abbreviations 2D: Two dimensional 3D: Three dimensional CDM: Cardiac differentiation medium CM: Cardiomyocyte cTnT: Cardiac muscle troponin T Cx43: Connexin 43 DEG: Differentially expressed gene ECM: Extracellular matrix EMT: Epithelial-mesenchymal transition Endo-MT: Endothelial-mesenchymal transition FA: Fatty acid FP: Field potential FPD: Field potential duration GSEA: Gene set enrichment analysis hESC: Human embryonic stem cell hiPSC: Human induced pluripotent stem cell hPSC: Human pluripotent stem cell MSC: Mesenchymal stem cell OCT: Optimal-cutting-temperature PC1: Principal component 1 PC2: Principal component 2 PFA: Paraformaldehyde SAA: Sarcomeric alpha-actinin TGF-β1: Transforming growth factor β1 α-SMA: α-smooth muscle actin References 1. 1. Sutra T, Oiry C, Azay-Milhau J, Youl E, Magous R, Teissedre PL, Cristol JP, Cros G. 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The molecular signatures database (MSigDB) hallmark gene set collection. Cell Syst. 2015;1(6):417–25. 79. 79. Jaffer S, Valasek P, Luke G, Batarfi M, Whalley BJ, Patel K. Characterisation of development and electrophysiological mechanisms underlying rhythmicity of the avian lymph heart. PLoS One. 2016;11(12):e0166428. Download references Acknowledgements Not applicable. Funding This work was supported by a grant from the Technology Innovation Program (No. 10063334) funded by the Ministry of Trade, Industry & Energy (MI, Korea), the National Research Foundation of Korea (NRF) grant funded by the Ministry of Science, ICT and Future Planning (2018R1C1B6008256 and NRF-2018M3A9H3023077), and the KRIBB Research Initiative Program. The funders had no role in the study design, data collection or analysis, decision to publish, or preparation of the manuscript. Availability of data and materials All data generated or analyzed during this study are included in this published article and its additional files. Author information MOL designed the studies, performed the experiments, analysed the data and prepared the manuscript. KBJ and JSJ performed the experiments and analysed the data. SAH, KSM, and JWS performed and analysed the MEA experiments. SHK and MYS planned the project, analysed the data, wrote and critically revised the manuscript. All authors read and approved the final manuscript. Correspondence to Mi-Ok Lee or Sang-Heon Kim or Mi-Young Son. Ethics declarations Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare that they have no competing interests. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Additional files Additional file 1: Movie 1. Cardiomyocytes derived from hESCs. (MP4 9007 kb) Additional file 2: Figure S1. Effect of TGF-β1 in CM spheroid. (A) Representative morphology of hESC-derived CM spheroids with or without TGF-β1 treatment for 3 weeks. (B) Masson’s Trichrome staining to visualize collagen fibers in CM spheroids. Scale bars, 100 μm. (MP4 4521 kb) Additional file 3: Movie 2. Cardiac spheroid of hESC-derived CMs on day 14. (MP4 4700 kb) Additional file 4: Movie 3. Cardiac spheroid comprising both hESC-derived CMs and MSCs on day 14. (MP4 1623 kb) Additional file 5: Movie 4. TGF-β1-treated cardiac spheroid comprising both hESC-derived CMs and MSCs on day 14. (MP4 1768 kb) Additional file 6: Figure S2. Collagen deposition in TGF-β1 treated CM-MSC microtissue. Masson’s Trichrome staining to visualize collagen fibres in multiple sections of CM spheroids at 14 days after 5 ng/ml TGF-β1 treatment. Scale bars, 100 μm. (TIF 5720 kb) Additional file 7: Figure S3. Comparative cellular component analysis of control and TGF-β1-induced fibrosis models. Gene set enrichment analysis (GSEA) of transcriptome data in TGF-β1 induced fibrosis model was performed by MSigDB of GO cellular component (580 gene set). (A) List of gene sets enriched in cardiac fibrosis model was shown by normalized enrichment score (NES) and false discovery rate (FDR). Enrichment plot of top ranked subset; proteinaceous extracellular matrix and basement membrane. (B) List of gene sets enriched in control was shown by NES and FDR value. Enrichment plot of top ranked subset, respiratory chain and inner mitochondrial membrane protein complex. (TIF 2203 kb) Additional file 8: Figure S4. Treatment of hESC-derived CMs with pro-fibrotic drugs. (A) Immunofluorescent staining of apoptotic CMs with an apoptosis-specific marker (Cleaved caspase 3; Cl-Casp3). Scale bars, 50 μm. Percentage of apoptotic CMs by quantifying ratio of Cl-Casp3 positive cells per number of DAPI-stained cells. C) Immunofluorescence staining of mitochondrial-specific marker (TOM20). Nuclei were stained with DAPI (blue). Scale bars, 10 μm. (TIF 5406 kb) Additional file 9: Table S1. List of the antibodies used in this study. (DOCX 16 kb) Additional file 10: Table S2. List of the primers used in this study. (DOCX 16 kb) Rights and permissions Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Reprints and Permissions About this article Verify currency and authenticity via CrossMark Cite this article Lee, M., Jung, K.B., Jo, S. et al. Modelling cardiac fibrosis using three-dimensional cardiac microtissues derived from human embryonic stem cells. J Biol Eng 13, 15 (2019) doi:10.1186/s13036-019-0139-6 Download citation Keywords • Cardiac fibrosis • Cardiac sphere • Cardiac microtissue • Cardiomyocyte • Mesenchymal stem cell
What Does Water Softener Do For Well Water If you are a resident of the Dallas-Forth Worth metroplex, you know that the water coming from your private well is corrosive in nature. The reason is the hard quality of water that enters your home from the main source. It contains excessive minerals including magnesium, calcium, lime and chalk. Ground water dissolves the rocks in the soil and acquires these minerals. As water from your well comes straight from the ground, it is not treated and hence contains such impurities. Water filters can help to a certain extent in making the water potable. However, when it comes to household chores, a larger quantity of water is required and it becomes difficult to purify it with the regular water filter. Why is hard water unfit for use? When hard water passes through your plumbing and fixtures, the minerals present in the hard water leave a layer of residue on clothes and dishes after wash. A thin white film is formed inside the bathtub and on the faucets which looks very ugly. If you wash your hair with such water, it loses its shine and feels sticky. Another problem that is generally faced because of hard water is that appliances do not work efficiently. When you use soap for bathing or detergent for washing, it does not form a rich lather because it reacts with calcium and magnesium in the water. This in turn leads to a build-up of soap which once again adds to the white and sticky layer. The soap scum is very stubborn and takes a long time to get removed. What is the best solution for this problem? The only way to prevent damage due to hard water is to soften the well water. A smart way of doing this is to install a special water filter which acts as a water softener. However, before installing a water softener, you should always test the water that comes out of your well for hardness. According to industry standards, if your water test result shows a figure of less than or equal to 1 GPG (grains per gallon) of calcium carbonate, your water is considered to be soft. It can be treated with simple water filtration. Anything above that level is considered either moderately or severely hard depending on the concentration of minerals. You can purchase a water analysis kit from companies that sell water softeners or you can call over technicians to carry out the testing process. All other methods of combating hard water except for water softening like descaling, filtration and chemical softening are either cost-prohibitive or ineffective in solving the problem completely. How does the Water softener work its magic on hard water? A water softener contains a resin tank along with a brine tank and a control system. The timer and control valve are connected to the water supply system inside your home. This device filters the hard water before it enters your property. First, the unconditioned water is allowed to flow into the resin tank which contains several plastic-like beads which are arranged at the bottom of the tank in the form of columns to form a resin-bed. This tank follows a simple principle. When the hard water flows over this resin bed, it switches the calcium and magnesium ions in the water with sodium ions or any other desirable ions. The resin beads are negatively charged and filled with sodium ions. These beads attract the positively charged ions of magnesium and calcium. Sodium has a weaker positive charge. Hence, magnesium and calcium ions with a stronger positive charge force the sodium ions to get displaced from the beads and mix with the water and they take its place instead. When the water reaches the bottom of the tank after penetrating the resin bed, it becomes clear of all impurities and is fit for use. This treated water is pumped out from the tank with the help of the riser tube and outlet manifold. After that, it is distributed throughout the house using the cold water pipeline. A high quantity of sodium is harmful for health but this process adds very little sodium to the water that enters your house. It does not pose any health concerns as such. You can safely use a water softener to get clean and pure water for daily use. How to keep the water softener running? The water softener is the most effective method of reducing the hardness of water. It works well for a long period of time till the calcium and magnesium ions replace all the sodium ions in the beads. Once that happens, the resin tank can no longer carry out the softening process effectively. It needs to be recharged with sodium periodically to function properly. This regeneration cycle involves loading up of the resin tank with a strong solution of salt and water from the brine tank. This backwash cycle involves a reversal of the flow of water as it is forced down the riser tube till it reaches the bottom of the tank. Then it is made to flow up the resin bed which leads to a flushing and expanding of the resin. The beads are washed and the extremely high amount of sodium chloride in this brine solution compels the magnesium and calcium ions to give away after which sodium takes their place in the beads once again. The brine solution containing the displaced ions of calcium and magnesium is flushed out through a drainpipe. After this process the brine tank is empty so a slow rinse starts with a gradual increase in speed. This flow of fresh water over the resin beads removes the extra salt from the beads and the tank and disposes it through the drain. The brine tank automatically gets refilled for the next regeneration cycle. Finally, the service cycle restarts and the resin tank is ready to soften the hard water again. There are various types of regenerating systems in water softeners. Some of them come with electronic timers which conduct regeneration at a set interval regularly. Some other softeners have a computer to calculate the extent of bead depletion on the basis of usage of water. Another system is that of the water meter which measures the amount of water used and starts the recharging process after the level of sodium gets exhausted. The system based on the amount of water processed is better than the timer-based one because it recharges only when necessary thereby, saving time and energy. Well Water Is Hard Water Benefits of using a water softener for treating well water: Removal of iron Along with calcium and magnesium, most water softeners also remove iron from your water supply, which is present in a high concentration in well water. Such a high level of iron is toxic for the body and can damage skin cells and cause damage to the stomach, liver, heart and pancreas. It can also stain the bathtub, sinks, dishes and showers along with clogging pipes. By installing the water softener, the well water entering your house is free of iron and hence, safe for use. Reduction of corrosion As the softening process removes the corrosive minerals from the water, it prevents further corrosion. There is no buildup of the residual minerals on different surfaces or on the inside of the pipes. This ensures de-clogging of the pipes and easy flow of water. The life of your plumbing system gets extended as the fixtures work effectively. Faster Heating of water Hard water takes longer to heat up than soft water. When you install a water softener, the water heater works 22% faster to heat the water. This improvement in efficiency reflects on your electricity bill as well by saving a lot of energy consumption. Healthier Drinking Water Everybody prefers drinking water which is free of impurities and clean because it is good for health. A water softener is able to remove most of the minerals from the water which are present in excess thereby improving the taste as well as the quality of the well water and making it completely fit for human consumption. This method is cheaper and safer than boiling water every time you need it for drinking or cooking. Lesser Buildup of Soap Scum As the concentration of minerals in the water decreases significantly, soap does not react with the water and turn into a sticky substance that is difficult to remove from the surface of the floor or sink. There is no white layer covering faucets, bathtubs and showerheads. This makes cleaning easier and reduces effort. You need to use less shampoo and soap while bathing because lather is easily formed and it cleans effectively. Along with these benefits, a water softener also preserves the life of appliances, keeps the dishes and fixtures clean and shiny, increases durability of fabrics and enhances smoothness of hair. Let VA Water LLC Help Make Hard Well Water Soft Now that you know how handy it is to have a water softener to treat well water, install one at your place today by getting in touch with VA Water LLC water filtration experts servicing the Dallas-Fort Worth-Arlington area!
Category: What is Differential Diagnosis? A differential diagnosis of a runny nose might consider hay fever as well as colds. Article Details • Written By: Mary McMahon • Edited By: Bronwyn Harris • Last Modified Date: 05 April 2014 • Copyright Protected: 2003-2014 Conjecture Corporation • Print this Article Free Widgets for your Site/Blog A 2003 blackout affected 50 million people in North America and had an economic impact of about $10 billion USD. more... April 23 , : William Shakespeare was born and died. (1564, 1616) more... In medicine, a differential diagnosis is a diagnosis which examines all of the possible causes for a set of symptoms in order to arrive at a diagnosis. For example, if a patient presents with a runny nose, doctors might consider causes like hay fever and colds in the differential diagnosis in an attempt to arrive at the right diagnosis. Many doctors use this system of diagnosis in their practices, and some doctors known as diagnosticians actually specialize in it. To perform a differential diagnosis, the physician starts by reviewing the patient's case. He or she interviews the patient to collect symptoms, and also gathers a family, personal, and social history which provides a picture of the patient's background. Typically, examinations and tests are also included, to get specific information about the patient's current condition. In a particularly puzzling case, the doctor might interview friends and coworkers or examine the patient's natural environment to look for causes. Once the physician collects all of the evidence, he or she considers possible causes for the patient's condition. Before any cause can be dismissed, the physician must come up with a good reason. Multiple causes are not ruled out in a differential diagnosis, as it is possible for patients to be sick with more than one thing at once. Once the physician has considered all the possible factors, he or she comes up with a working diagnosis and begins treatment. Ad The patient's condition under treatment is monitored, and if the patient fails to improve or gets worse, the doctor returns to the drawing board. The failure of treatment is incorporated into the differential diagnosis, and the doctor begins again. In a sense, this type of diagnosis is a process of elimination, but it can get extremely involved and very complex, since many diseases and common afflictions look very similar, and it is easy for a physician to be misled. Working out a diagnosis using this method can be like assembling the pieces of a puzzle, especially when a patient has conflict symptoms or a complex history. It is an important part of the practice of internal medicine, a medical specialty which focuses on the diagnosis and treatment of nonsurgical conditions in patients. Internists are sometimes called in to assist other medical specialists with a differential diagnosis, since they are able to draw upon a wide body of knowledge and experience. Ad Discuss this Article cafe41 Post 3 @Cupcake15 - I think that that is the hardest part of medicine because you have to do a lot of research and even though you have a lot of information, you still may not know what the patient has which has to be really disheartening. I remember watching a television program that dealt with this sort of thing. The patients all had progressively worsening conditions, but the doctors continually misdiagnosed the problems and only after the patients sought renowned specialists did they get an accurate medical differential diagnosis. There was one case where a little girl would get bruised just by playing on the floor and one day she was bleeding uncontrollably. It was eventually discovered that the little girl had a very rare disease that did not allow her blood to clot correctly which is why with the littlest scratch she would make her bleed. The doctors were able to give the little girl the right medication and she finally received some relief from the disorder which was also a big relief for her family as well. cupcake15 Post 2 @Sunny27 - I agree and I have to say that conditions that result in chest pains or abdominal pain are very serious and every possibility should be considered. For example, an abdominal pain differential diagnosis should also consider if there is any bleeding associated with the pain which could mean that the patient may have an ulcer or may have something as serious as cancer. There is such a wide spectrum of potential ailments that the differential diagnosis pathology has to include a number of tests and reviews of the family history in order to reveal what the patient is suffering from. Sunny27 Post 1 I think that a medical differential diagnosis is important because a lot of disorders have similar symptoms and it is important to explore every possibility before ruling out any condition. For example, a lot of women experience chest pain and while some doctors will offer a chest pain differential diagnosis to rule out heart disease many doctors don’t because heart disease is misdiagnosed with women. They say that more women die of heart attacks than men because doctors mistakenly confuse the symptoms with something else. Post your comments Post Anonymously Login username password forgot password? Register username password confirm email
Select Page Track bearings bearing replacement Track Bearings Bearing Replacement Introduction Track bearings play a crucial role in various applications, providing smooth and precise movement in industrial machinery. Over time, these bearings may wear out and require replacement. In this article, we will explore the process of track bearings bearing replacement, discussing the necessary steps and considerations to ensure a successful replacement. 1. Understanding Track Bearings In order to effectively replace track bearings, it is important to have a clear understanding of their construction and function. Track bearings are designed to support and guide rotating shafts or axles, allowing them to move smoothly along a track or rail. They consist of an outer race, an inner race, rolling elements (such as balls or rollers), and a cage that holds the rolling elements in place. 2. Signs of Worn-out Track Bearings Before proceeding with the replacement, it is crucial to identify the signs of worn-out track bearings. Some common indicators include: • Unusual noises or vibrations during operation • Inconsistent or jerky movement • Increased friction or resistance • Visible damage or wear on the bearing surfaces If any of these signs are observed, it is recommended to replace the track bearings promptly to prevent further damage to the machinery. 3. Steps for Track Bearings Bearing Replacement Replacing track bearings requires careful planning and execution. Here are the essential steps to follow: 1. Prepare the necessary tools and equipment, including a suitable replacement track bearing and any specialized tools required for installation. 2. Ensure the machinery is properly shut down and isolated from any power source. 3. Remove any protective covers or guards that may be obstructing access to the track bearing. 4. Using the appropriate tools, carefully remove the old track bearing from its housing, taking care not to damage any surrounding components. 5. Clean the housing and surrounding area thoroughly to remove any debris or contaminants. 6. Inspect the housing for any signs of damage or wear. If necessary, repair or replace the housing before proceeding with the installation. 7. Apply a suitable lubricant to the replacement track bearing to ensure smooth operation. 8. Insert the replacement track bearing into the housing, aligning it properly with the shaft or axle. 9. Secure the track bearing in place using the appropriate fasteners or locking mechanisms. 10. Reinstall any protective covers or guards that were removed earlier. 11. Test the machinery to ensure the new track bearing is functioning correctly and providing the desired movement. Conclusion Bearing replacement is an essential maintenance task to ensure the optimal performance and longevity of track bearings. By following the proper steps and guidelines, you can successfully replace track bearings and restore the smooth and precise movement in your industrial machinery. About the Author Author: Czh Track Bearings Application Company Introduction We are a leading company in the Chinese reducer market, specializing in the production of various high-quality products. Our range includes servo reducers, plastic gearboxes, gear motors, worm gearboxes, worm wheels, and worm reducers. With state-of-the-art CNC production and assembly equipment, we ensure the highest standards of quality and precision in our products. At our company, we pride ourselves on offering competitive prices and exceptional customer service. We welcome customization requests from our customers, providing tailored solutions to meet their specific requirements. Choose our products for superior performance, attractive pricing, and reliable service. Company Factory Author: Czh Tags: SSJ GROUP PTE. LTD. As one of the leading manufacturers, suppliers, and exporters of mechanical products, We offer reducers, sprockets, industrial and conveyor chains, belts, pulleys, gears, racks, gearboxes, motors, PTO Shafts, taper lock Bushing, and many other products. Please get in touch with us for details. SSJ UK Limited Sara Jones Mail: [email protected] TEL: + 44 7944 869861 ADDR.: Bury St Edmunds, Suffolk IP32 7LX, UK SSJ ITALY S.R.L. TEL: +39 0522 1606 388; +39 3471 65 17 22 ADDR.:Via Pasteur, Reggio Emilia, Italy Recent Updated
Root - Orange San Francisco 2 (ZTE Blade 2) Learn how to root the Orange San Francisco 2 (ZTE Blade 2) download ROMs and kernels, discuss recovery options, and get help with everything related to Orange San Francisco 2 (ZTE Blade 2) rooting. Last Updated: Sort by: Title Replies Views Last Message ↓ 1. agentc13 Replies: 0 Views: 12,286 2. » 3. Sanfrancisno Replies: 1 Views: 482 4. Sanfrancisno Replies: 0 Views: 838 5. christophe195 Replies: 5 Views: 1,152 6. silent785 Replies: 4 Views: 2,919 7. rocker123 Replies: 20 Views: 6,352 8. rocker123 Replies: 0 Views: 1,538 9. MustangV10 Replies: 1 Views: 838 10. Bencaramax Replies: 1 Views: 3,604 11. fuzzface Replies: 4 Views: 5,007 Thread Display Options Loading...
back_image blog-image How Can You Create Your Own Flutter App for WordPress? An in-depth Guide Author Image By Editorial Team Last Updated: August 26, 2024 6 minutes 00:00:00 Reading time: 0s The goal of every website is to be effortlessly navigable across several platforms, including desktop, mobile, web, and iOS. Since it offers a smooth and efficient experience to the customer, which accurately reflects the brand and enhances the goodwill in the market. However, From the developers’ point of view, creating a flutter app for a WordPress website is a challenging task. Therefore, here we’ll walk you through the simple and straightforward process of building a Flutter app within a few steps, along with its benefits. Here’s How You Can Create a Flutter App for Your WordPress Website How to Create a Flutter App for WordPress Following these 6 steps, you can create your Fluter app and run it on the WordPress website smoothly. So, let’s follow them! Step 1: Step up your Flutter Environment Here, first, you have to install the Flutter framework by following the official installation instructions exclusively. After it’s finished, you need to choose an IDE for developing Flutter apps, such as Android Studio or Visual Studio Code. Step 2: Create a New design for the App Next to that, you have to generate a new Flutter project using the Flutter CLI (Flutter Command Line). This will assist in setting up a web directory inside your project and enable Flutter for the web. Step 3: Assemble the user interface In this step, you have to set up the UI (User Interface) for your Flutter application using the Flutter widgets. From here, you can use HTTP queries to the WordPress REST API to retrieve the data from your WordPress website. Step 4: Display Your WordPress-Generated Stuff Now, you need to get the material from your WordPress website and have to use it in your Flutter application by filling it out in the user interface space. Step 5: Test your App For better testing, you are required to test your app on both web browsers and mobile emulators or devices. Make sure the UI and style of your app are mobile-friendly. Step 6: Publishing the App At last, you need to be ready to launch your Flutter app on the Apple App Store and Google Play Store. Note:- Many people don’t want to use placeholder posts in their WordPress framework while creating their fluter application whereas, some love to do so. Therefore, let me assist you in figuring out whether you need to use the placeholder post in your WordPress or not. Advice: Instead of using a placeholder post, you can move towards WordPress plugins that will not only handle your workflow smoothly but also will offer site security, better SEO, e-commerce functionality & more. Why do you need to Create a Flutter App for Any WordPress Site? Creating a flutter application for any WordPress website not only generates the strong and professional goodwill of the brand but also provides the following benefits! 1. Expressive and Flexibe UI: Using the WordPress framework while creating the Flutter app, you can have the experience of a rich set of pre-designed widgets for creating customized UIs. 2. Smooth Performance: Here, you are allowed to use the Dart programming language and compile it to native code, ensuring high performance. 3. Single Codebase: If you are designing a flutter app using the WordPress framework then you have to write a single code once and you can run it on multiple platforms further. 4. Strong Community Support: It provides large community support and an extensive library of packages along with plugins 5. Integration with Existing Code: Flutter can easily get integrated with existing web applications and can coexist with other technologies. Consequently, you’re allowed to incrementally migrate to Flutter without having to rewrite the entire codebase. 6. Quick Development: The hot reload feature will allow you to make quick changes and instant feedback together. 7. Access to Native Functionality: Experiencing Flutter, you can get free access to the native device features such as camera, GPS, and storage as flutter provides these features. You can use this for creating web applications that require access to device-specific functionalities. 8. Open Source Service: As all know, flutter is an open-source framework, you can use it for free and get a transparent development process. Plus, you can customize it accordingly to meet your specific needs. Here you have got the reasons that will push you towards using the WordPress framework while designing your Flutter application. Therefore, let’s have a look at the mentioned points to know the stepwise guidance on creating a flutter app for any WordPress website. What is a Paceholder Post? A placeholder post refers to a post, image, or dummy text which is used as a temporary entry on a website or blog to reserve a spot for future content. For Example:- Generic Titles, Sample Images, Categories and Tags, Dummy Text, and so on. Basically, it contains minimal information such as a title, basic text, or a “coming soon” message. Website developers use placeholder posts to maintain a consistent structure on the site and allow easier planning and organization of upcoming content. Placeholder posts are particularly useful during the design and development phase to ensure that the layout and navigation elements functioning correctly even when the final content is not yet available or not. Once the actual content is ready then the developers replace the placeholder post with the full post. Why Should You Remove Placeholder Posts in WordPress? When website developers create a Flutter app that integrates with a WordPress backend then it is essential to know how to get rid of placeholder posts for several reasons: 1. Data Integrity: Placeholder posts give a cluttered look to the database and make it harder to manage and retrieve the real content. So, avoiding placeholder posts can keep your database clean and ensure that all data is valid and relevant. 2. Performance: If you will use excessive placeholder content in your website development then it can slow down the app by increasing the amount of data that needs to be loaded and processed. Plus, eliminating the placeholder posts can enhance the performance and responsiveness of your application. 3. SEO and Analytics: If you will use placeholder posts it will impact your SEO negatively by creating irrelevant or duplicate content. In addition to this, it will also skew analytics data, making it difficult to track user engagement and behavior accurately. 4. User Experience: The biggest reason behind not using placeholder posts is that placeholder posts with dummy text and images can confuse users and make your app look unprofessional. Additionally, removing them ensures that only relevant and meaningful content is presented to users. 5. Content Accuracy: Typically the placeholder posts get indexed by search engines or shared by users which leads to misinformation or outdated content being circulated. So, you need to sure that your entire content is accurate and up-to-date to avoid further issues. 6. Professional Appearance: If you will use a clean and placeholder-free app, then it will appear more polished and trustworthy to users. Which will be useful in building credibility and a positive impression of your application. 7. Maintenance: In case you have decided to move to using placeholder posts, don’t forget to be ready to manage a website or app with additional effort to ensure the placeholders don’t go live accidentally. Consequently, removing placeholder posts from your development reduces the risk of such mistakes and simplifies content management. 8. Consistency: Placeholder posts can disrupt the consistency of your application’s content. So, you need to remove them to ensure a uniform presentation and coherent user experience. Let’s Recap Before You Go! This blog post will give you a comprehensive guide on creating a flutter app for any WordPress website in just only 5 steps along with the proper behind-the-scenes causes. Moreover, you will get a complete knowledge of the placeholder posts, their role, their essentiality in the WordPress framework, and the steps to remove them. So, without making any delay let’s jump into the main content. Categories: Technology 0 Comments Leave a Reply Avatar placeholder
Skip to main content Advertisement We’d like to understand how you use our websites in order to improve them. Register your interest. A role for cathepsin Z in neuroinflammation provides mechanistic support for an epigenetic risk factor in multiple sclerosis Abstract Background Hypomethylation of the cathepsin Z locus has been proposed as an epigenetic risk factor for multiple sclerosis (MS). Cathepsin Z is a unique lysosomal cysteine cathepsin expressed primarily by antigen presenting cells. While cathepsin Z expression has been associated with neuroinflammatory disorders, a role for cathepsin Z in mediating neuroinflammation has not been previously established. Methods Experimental autoimmune encephalomyelitis (EAE) was induced in both wildtype mice and mice deficient in cathepsin Z. The effects of cathepsin Z-deficiency on the processing and presentation of the autoantigen myelin oligodendrocyte glycoprotein, and on the production of IL-1β and IL-18 were determined in vitro from cells derived from wildtype and cathepsin Z-deficient mice. The effects of cathepsin Z-deficiency on CD4+ T cell activation, migration, and infiltration to the CNS were determined in vivo. Statistical analyses of parametric data were performed by one-way ANOVA followed by Tukey post-hoc tests, or by an unpaired Student’s t test. EAE clinical scoring was analyzed using the Mann–Whitney U test. Results We showed that mice deficient in cathepsin Z have reduced neuroinflammation and dramatically lowered circulating levels of IL-1β during EAE. Deficiency in cathepsin Z did not impact either the processing or the presentation of MOG, or MOG- specific CD4+ T cell activation and trafficking. Consistently, we found that cathepsin Z-deficiency reduced the efficiency of antigen presenting cells to secrete IL-1β, which in turn reduced the ability of mice to generate Th17 responses—critical steps in the pathogenesis of EAE and MS. Conclusion Together, these data support a novel role for cathepsin Z in the propagation of IL-1β-driven neuroinflammation. Introduction Enigmatic to the pathogenesis of multiple sclerosis (MS) are the mechanisms that link known risk factors to the incidence and development of this immune-driven demyelinating disease. Since the incidence of MS is influenced by environmental factors and gender, but has a low concordance rate in monozygotic twins and underwhelming odds ratios for individual SNPs, epigenetic changes are likely to play a major role in determining an individual’s susceptibility to MS [1]. In 2014, Huynh et al. compared epigenomic differences between pathology-free regions of healthy and MS-affected brains in an attempt to identify potential epigenetic risk factors for MS [2]. One of the most significant findings was that the cathepsin Z (CTSZ) locus was hypomethylated in pathology-free regions of MS patients, which resulted in increased expression of cathepsin Z within this neural tissue [2]. While the underlying mechanism that results in hypomethylation at this particular locus is unknown, the authors proposed that the epigenetically-driven expression of cathepsin Z in neural tissue may increase an individual’s susceptibility to MS. Cathepsin Z (also known as cathepsin X) was identified in silico by its similarity to the family of cysteine-type lysosomal proteases, through mining the Expressed Sequence Tags database from human brain tissue [3, 4]. Cathepsin Z is a unique member of this 11 member-protease family, as it is the only enzyme with strict carboxypeptidase activity, it has a remarkably short pro-domain that contains a RGD integrin binding domain, and the CTSZ gene is chromosomally separated from the other lysosomal cysteine cathepsin genes [5,6,7,8]. In the context of neoplasia, there is recent in vivo evidence for a tumor-promoting role for the carboxypeptidase as well as for the RGD function of cathepsin Z [9]. However, to date, the specific functions of cathepsin Z within the central nervous system (CNS) remain obscure. Whilst there was mounting evidence to support the association of cathepsin Z expression with neuroinflammation [10,11,12,13], whether cathepsin Z had a specific pathogenic role in neuroinflammatory disorders was erstwhile unknown. Here, we present experimental evidence to support a non-redundant role for cathepsin Z in neuroinflammation in mice. In a model of multiple sclerosis—experimental autoimmune encephalomyelitis (EAE)—mice deficient in cathepsin Z consistently developed lower levels of neuroinflammation and displayed disproportionally lower levels of circulating IL-1β. The ability to generate IL-1β in response to NLRP3-stimulus by macrophages and dendritic cells derived from cathepsin Z-deficient mice was compromised, as was the ability of cathepsin Z-deficient mice to generate Th17 responses. Collectively, these data indicate that cathepsin Z promotes the IL-1β–Th17 axis leading to more severe neuroinflammation during EAE in mice and may suggest a role for cathepsin Z in the development of MS, as proposed by Huynh et al. [2]. Materials and methods Mice and cells C57BL/6 (wildtype [WT]) and C57BL/6-Tg(Tcra2D2,Tcrb2D2)1Kuch/J (2D2) mice were purchased from the Jackson Laboratory (Bar Harbor, ME, USA). 2D2 mice express a transgenic CD4+ T cell receptor (Vβ11 TCR/Vα3.2 TCR) that is specific for the immunodominant MOG35–55 peptide in the context of I-Ab [14]. Cathepsin Z-deficient mice (Cat Z−/−) were generated as previously described [15]. In brief, a segment of the murine cathepsin Z exon 2, containing a portion of the active site along part of intron 3, was substituted with a ribosomal entry sequence [15]. Cathepsin Z-deficient mice were also crossed with 2D2 mice, generating mice with cathepsin Z-deficient 2D2 CD4+ T cells. All mice used were fully backcrossed to the C57BL/6 background, and bred and housed under identical animal husbandry conditions. All animal research was performed in accordance with the Canadian Council for Animal Care, and protocols were approved by the University of Calgary Animal Care and Use committee. All mice were age and sex matched within experiments, and used between the ages of 8 and 12 weeks. All mice subject to in vivo studies were 8–10 weeks of age [16, 17]. Bone marrow-derived macrophages (BMMØs) were derived from bone marrow using L929-conditioned media and activated for 18 h with recombinant IFNγ (rIFNγ) (100 U/ml, Pepro Tech), as previously described [18, 19]. Bone marrow-derived dendritic cells (BMDCs) were derived using media conditioned with the supernatant of Ag8.653 myeloma cells transfected with GM-CSF cDNA, as previously described [16, 20,21,22]. Peritoneal macrophages (pMØs) were isolated from the peritonea of mice by injection of 8 mL of sterile PBS using a 23 g needle, followed by removal of 5 ml of PBS/peritoneal fluid after brief abdominal massage [23]. The murine microglia-like cell line BV2 (C8-B4 [ATCC® CRL-2540™]) was grown in RPMI supplemented with 5% FBS. The murine albino neuroblastoma cell line Neuro-2a (N2A) (ATCC® CCL-131™) was grown in DMEM/F-12 supplemented with 5% FBS. The murine dendritic cell-like DC2.4 (CVCL_J409) was grown in RPMI supplemented with 5% FBS, 10 mM β-mercaptoethanol (2-ME), 20 mM L-glutamine and 100 mM HEPES [24]. All cells were cultured at 37 °C with 5-7% CO2. Flow cytometry Flow cytometry was performed using a FACSCalibur flow cytometer (BD Biosciences, Franklin Lakes, NJ, USA) and analyzed with FLOWJO software v8.6 (Tree Star, Ashland, OR, USA) [16]. Leukocyte populations, with a minimum of 2.5 × 104 counts, were selected using forward scatter/side scatter (FSC/SSC). All antibodies were purchased from BD Biosciences [16]. Assessment of MOG antigen processing and presentation MOG antigen presentation efficiency was assessed by measuring the relative level of activation of CD4+ T cells derived from the TCR transgenic mouse model 2D2 following co-culture with MOG-pulsed antigen presenting cells (APCs), as previously described [14, 16, 25,26,27]. In brief, WT and Cat Z−/− BMMØs and BMDCs were exposed for 6 h to medium containing the immunodominant I-Ab peptide epitope MOG35–55 (0, 1, 10, 25 μg/ml), synthesized by the University of Calgary Peptide Services (AB, Canada), or the full extracellular domain of MOG (MOG1–125; 0, 1, 10, 25 μg/ml), prepared as previously described [16]. APCs were then washed with T cell media (RPMI supplemented with 10% FBS and 10 mM 2-ME), and naïve 2D2 or Cat Z−/− 2D2 splenocytes were added to the APCs and incubated for 16 h [16]. The presentation efficiency of MOG35–55 by the WT and Cat Z−/− APCs, to the 2D2 or the Cat Z−/− 2D2 CD4+ T cells, was determined cytometrically using the surface expression of the early activation marker, CD69. CD4+ T cell trafficking CD4+ T cell trafficking was assessed as previously described [28]. In brief, WT and Cat Z−/− CD4+ T cells were isolated from spleens of WT and Cat Z−/− mice using the EasySep Mouse CD4+ T Cell Enrichment Kit (StemCell Technologies), according to the manufacturer’s instructions. 3 × 105 CD4 + T cells were transferred to the upper filter of a 5 μm Transwell support plate (Corning) pre-coated overnight with 3 μg/ml ICAM-Fc (R&D Systems). T cells were allowed to settle for 30 minutes before the upper filter was exposed to the bottom chamber containing vehicle (PBS) or 1 μg/ml CXCL9 (R&D Systems), followed by an incubation period of 1 h at 37 °C. The absolute numbers of cells that migrated through the Transwell filter were enumerated with a hemocytometer. Induction of EAE and adoptive transfer of 2D2 CD4+ T cells EAE was induced in WT and Cat Z−/− mice using standard protocols, as previously described [16, 17]. In brief, 8- to 10-week-old female WT and Cat Z−/− mice were anesthetized using intraperitoneal (i.p.) injection of ketamine-xylazine. Following anesthetization, each mouse was injected sub cutaneously (s.c.), in both flanks, with a 100 μL emulsion of 50 μg MOG35–55 in complete Freund’s adjuvant (0.5 mg/ml M. butyricum in paraffin oil) (CFA; BD). Additionally, each mouse received an i.p. injection of pertussis toxin (PT) (300 ng) on days 0 and 2, and the clinical score and weight was recorded daily for the duration of the experiment. The following clinical scoring system was used: score 0 - asymptomatic; 0.5 - tail weakness; 1 - limp tail; 1.5 - hind limb limping; 2 - hind limb weakness; 2.5 - partial hind limb paralysis; 3 - complete hind limb paralysis; 3.5 – hind limb paralysis with forelimb weakness; 4 – forelimb paralysis; 4.5/5 - complete morbidity/death [16, 17]. Mice that received saline/CFA/PT did not develop clinical signs of EAE (data not shown). Additional cohorts of mice were sacrificed at day 15 for CNS leukocyte analysis and cardiac puncture for Luminex analysis of peripheral cytokines [16]. To investigate the ability of WT and Cat Z−/− mice to generate Th17, Th1 and Treg CD4+ T cell responses in the absence of MOG, anesthetized mice were injected s.c., in both flanks, with a 100 μL saline/CFA emulsion. These mice were sacrificed 6 days after injection, the inguinal lymph node cells (LNC) were removed and CD4+ T cells expressing IL-17, IFNγ, and FoxP3 were counted by flow cytometry. To examine CD4+ T cell migration to and activation within the CNS, 2D2 and Cat Z−/− 2D2 CD4+ T cells were activated, expanded, and adoptively transferred into WT or Cat Z−/− mice, as previously described [29]. Briefly, 2D2 and Cat Z−/− 2D2 splenocytes were harvested and cultured in T cell medium (RPMI supplemented with 10% FBS and 10 mM 2-ME) containing 20 μg/ml MOG35–55 and 0.5 ng/ml IL-12 (R&D Systems) for 48 h. The expanded 2D2 or Cat Z−/− 2D2 CD4+ T cells were injected into the peritoneal cavities of WT or Cat Z−/− mice (5 × 106 total cells/mouse suspended in 100 μL PBS) [16]. Adoptively transferred CD4+ T cells were isolated 6 days later using a discontinuous Percoll gradient, immunostained for CD4+, Vα3.2+ (2D2 TCR) and CD25, and analyzed by flow cytometry. Spinal cord leukocyte profiling Infiltrating and resident leukocytes of the spinal cord were isolated 15 days post EAE induction using a discontinuous Percoll gradient, as previously described [16, 30]. These cells were immunostained in the following combinations: macrophages (CD11b+/CD45+ high), B cells (B220+/CD45+), CD8+ T cells (CD8+/CD3+), CD4+ T cells (CD4+/CD3+) and Th17 cells (IL-17+/CD4+/CD3+), and analyzed by flow cytometry. Histology The thoracolumbar spinal cord of each WT and Cat Z−/− mouse, sacrificed at day 15 after EAE induction, was removed in toto and fixed in 10% neutral buffered formalin. Transverse sections of the lumbosacral spinal cord from the lumbar intumescence were paraffin embedded, sectioned at 3 μm and stained with hematoxylin and eosin (HE) and Luxol fast blue (LFB) (Prairie Diagnostic Services, Saskatoon, SK, Canada). Fluorometric assessment of phagolysosomal proteolysis Proteolytic efficiencies of phagolysosomes in live phagocytes were measured, as previously described [18, 19]. In brief, WT and Cat Z−/− BMDC and BMMØs were allowed to phagocytose 3 μm silica beads that were covalently coupled to human IgG (Sigma) and to the fluorogenic protease substrate DQ Green BODIPY albumin (DQ-albumin; Invitrogen) bearing the reference fluor (Alexa Fluor 594 succinimidyl ester; Invitrogen). The proteolytic efficiencies of the resulting phagolysosomes were determined by measuring the green fluorescence liberated from hydrolysis of the particle-bound DQ-albumin, relative to that of the reference fluor, over a one-hour period. Measurements were performed at 37 °C in microplate format using an Envision multilabel plate reader (PerkinElmer Life Sciences). qPCR Quantitative PCR (qPCR) was used to quantify key mRNA transcripts in spinal cord tissue over the course of EAE, as well as in WT and Cat Z−/− APCs in response to LPS [16]. In brief, WT lumbar spinal cords, spleens, neurons or APCs (including pMØs, DC2.4 s, BV2s, BMMØs and BMDCs treated with LPS for 3 h) were snap-frozen in liquid nitrogen, total RNA was extracted using the RNeasy lipid tissue mini kit (Qiagen), and cDNA was synthesized using iScript Reverse Transcriptase Supermix for RT-qPCR (BioRad). qPCR was performed as previously described [19]. All primers were prepared at 300 nM, had a single melt curve, had efficiencies between 90–100%, and were designed or verified using Primer 3 (National Center for Biotechnology Information). 18S (F: 5′- AGTCGGCATCGTTTATGGTC-3′; R: 5′-CGCGGTTCTATTTTGTTGGT-3′) was used as an internal control, and did not vary across treatments; IL-1β (F, 5′-CAACCAACAAGTGATATTCTCCATG-3′; R, 5′-GATCCACACTCTCCAGCTGCA-3′) and Cat Z (F, 5′- CCTGTCCGGGAGGGAGAA −3′; R, 5′- TGGTTGATAACGGCCTGGTC −3′) [31] were amplified using the following PCR conditions (in a BioRad iQ5 thermocycler): 95 °C for 5 min; 40 cycles of 95 °C for 30s and 55 °C (58 °C for 18S and IL-1β) for 30s. All mRNA levels were presented relative to 18S and the WT control samples. IL-1β and IL-18 generation by APCs APC generation of IL-1β and IL-18 in response to activation of the NLRP3 inflammasome in vitro were quantified by Mouse IL-1β ELISA and Mouse IL-18 Platinum ELISA (eBioscience) [32]. In brief, WT and Cat Z−/− BMMØs, and BMDCs were pre-incubated with 200 ng/ml of ultra-pure LPS from Salmonella minnesota R595 (List biological laboratories) for 3 h. Following a single wash with warm PBS, BMDCs were incubated with adenosine triphosphate (ATP 5 mM) for 1 h (Sigma-Aldrich); and BMMØs with monosodium urate crystals (MSU, 300 ng/ml prepared as in [33]) for 6 h in appropriate media. Supernatants were collected and protein concentrations were measured by ELISA according to the manufacturer’s instructions. Statistical analysis Statistical analyses were performed by one-way ANOVA with a Tukey post hoc test, or an unpaired Student’s t test, as specified (p < 0.05). If a Bartlett’s test for equal variance failed, then the data underwent a natural log transformation before reanalysis. Analyses of EAE clinical scoring were performed using the non-parametric Mann–Whitney U test [16]. Analyses were completed using GraphPad Prism software (La Jolla, CA, USA). Results Cathepsin Z does not significantly impact proteolytic efficiencies within phagolysosomes of macrophages The lysosomal cysteine protease, cathepsin Z, has been reported to be highly expressed in antigen presenting cells (APCs) [12, 13, 34]. This was confirmed in peritoneal and bone marrow-derived macrophages (pMØ, BMMØ), immortalized and bone marrow-derived dendritic cells (DC2.4, BMDC) and immortalized microglia (BV2) (Fig. 1a). Although the functions of cathepsin Z are ill-defined, it is well accepted that the primary function of many lysosomal cysteine cathepsins—such as cathepsin B, L and S—is to facilitate protein turnover by hydrolyzing self and foreign proteins in the cellular endolysosomal network. This system is particularly well developed in phagocytic APCs such as macrophages and dendritic cells, where proteolytic cleavage by these enzymes within phagosomes and endosomes is also necessary for the processing of T cell antigens [35,36,37]. In order to determine the relative contribution of cathepsin Z to endolysosomal proteolysis, the hydrolysis of phagocytosed protein was measured in WT and Cat Z−/− BMMØs and BMDCs. Unlike specific deficiencies in other lysosomal cysteine cathepsins (such as cathepsin S), BMMØs and BMDCs derived from mice deficient in cathepsin Z displayed comparable efficiencies of phagosomal proteolysis (Fig. 1b–e). Fig. 1 figure1 Cathepsin Z is highly expressed in APCs but does not significantly contribute to phagolysosomal proteolysis. a Cathepsin Z mRNA levels in BV2 (C57BL/6 brain microglia cell line), DC2.4 (dendritic cell line), BMDC (bone marrow derived dendritic cells), pMØs (peritoneal macrophages), BMMØ (bone marrow derived macrophages), N2A (murine albino neuroblastoma cell line) and Cat Z−/− BMDCs (n = 3). b-e The total proteolytic activity (rate of substrate-liberated fluorescence from the particle-bound fluorogenic substrate DQ-albumin) following phagocytosis of fluorometric experimental particles in WT and Cat Z−/− (b-c) BMMØ (n = 9) and (d-e) BMDC (n = 5). b, d Representative real-time traces of phagosomal proteolysis. c, e Averaged rates of proteolysis (determined by calculation of the slope of the linear portion of the real-time trace [as described by y = mx + c, where y = relative fluorescence, m = slope, and x = time] were calculated between (c) 20 min and 60 min or (e) 20 min and 40 min after particle internalization. Data presented as mean+/− SEM; (c, e) no significant differences (unpaired Student’s t-test, p > 0.05) from the WT control were observed Mice deficient in cathepsin Z show attenuated neuroinflammation during EAE In concordance with a previous microarray study [38], cathepsin Z mRNA was found to be significantly upregulated in neural tissue during murine EAE following induction with myelin oligodendrocyte glycoprotein peptide (MOG35–55) using qPCR (Fig. 2a). To determine whether cathepsin Z plays an active role in the pathogenesis of EAE or is merely an indicator of neuroinflammation, several parameters of neuroinflammation were compared between C57BL/6 (WT) and congenic cathepsin Z-deficient (Cat Z−/−) mice following the induction of EAE. Consistent with a potential role for cathepsin Z in neuroinflammation, Cat Z−/− mice displayed significantly attenuated progression of the clinical signs of EAE compared to that of WT mice (Fig. 2b, Additional file 1: Figure S1). At 15 days post induction (EAE peak), spinal cord tissue in Cat Z−/− mice showed reduced demyelination and neuroinflammation by histopathology, as well as diminished infiltration of macrophages and lymphocytes, as determined by flow cytometry (Fig. 2c, d). Fig. 2 figure2 Cathepsin Z expression is increased in the CNS during EAE, and mice deficient in cathepsin Z exhibit attenuated signs of neuroinflammation and demyelination during EAE. a Cathepsin Z mRNA levels in the spinal cord tissue of WT mice 15 days after induction of EAE or mock (n = 6). b Clinical disease course of WT and Cat Z−/− mice after active induction of EAE (n = 20–21). c Representative micrographs of transverse sections of lumbar spinal cord from WT and Cat Z−/− mice at 15 days after induction of EAE. Sections are stained with hematoxylin and eosin (HE) for inflammation, or Luxol fast blue (LFB) for demyelination. Grey and black scale bars indicate 500 and 100 μm respectively. d The total number of infiltrating macrophages (MO), B cells, CD8+ T cells (CD8+), CD4+ T cells (CD4+), and Th17 cells (IL-17+/CD4+) isolated from lumbar spinal cord tissue 15 days post EAE induction as analyzed by flow cytometry (n = 8–12). Data presented as mean +/− SEM; significant differences (unpaired Student’s t test; clinical data, Mann–Whitney U test; p < 0.05) from the WT control are denoted by asterisks Cathepsin Z deficiency does not impact either the processing and presentation of the autoantigen MOG or CD4+ T cell activation and trafficking To determine whether cathepsin Z-deficiency impacts EAE through perturbation of the processing and/or presentation of the autoantigen MOG, APCs derived from WT and Cat Z−/− mice were incubated with pre-processed MOG peptide (MOG35–55) or unprocessed MOG protein (MOG1–125) and co-cultured with MOG35–55-specific CD4+ T cells. Both BMMØs and BMDCs from Cat Z−/− mice were able to activate and mature in response to proinflammatory stimuli and could efficiently activate MOG35–55-specific CD4+ T cells in an antigen-specific fashion (as determined by expression of the early T cell activation marker, CD69) (Fig. 3a–d). Consistent with an insignificant role for cathepsin Z in phagosomal proteolysis, these data demonstrate that cathepsin Z does not impact the processing or presentation of the autoantigen MOG, suggesting that its role in the pathogenesis of murine EAE is mediated through a mechanism not previously attributed to lysosomal cysteine cathepsins [39]. To investigate a potential role of cathepsin Z in T cell functions germane to EAE, Cat Z−/− CD4+ T cells were evaluated for their ability to activate, chemotax and respond to MOG. It was found that CD4+ T cells isolated from WT and Cat Z−/− mice were equally able to activate in a MOG35–55-specific fashion (Fig. 3e). Similarly, the absence of cathepsin Z did not affect the ability of CD4+ T cells to chemotax in response to CXCL9 (Fig. 4a). Consistent with these findings, adoptively transferred WT and Cat Z−/− MOG35–55-specific CD4+ T cells traversed the blood-brain-barrier and responded to the presence of endogenous MOG within the CNS in comparable fashions (Fig. 4b, c). Fig. 3 figure3 Cathepsin Z-deficiency does not affect the efficiency of MOG antigen processing or presentation in BMMØ and BMDC, or the efficiency of MOG-specific CD4+ T cell activation. a-e WT and Cat Z−/− (a, b, e) BMMØ and (c-d) BMDC were incubated with (a, c, e) MOG35–55 peptide (0, 1, 10, 25 μg/ml), or (b, d) recombinant MOG1–125 (0, 1, 10, 25 μg/ml) for 6 h before co-incubation with (a-d) MOG35–55-specific 2D2 CD4+ T cells or (e) WT or Cat Z−/− MOG35–55-specific 2D2 CD4+ T cells. Activation of WT and Cat Z−/− MOG35–55-specific 2D2 CD4+ T cells was assessed via CD4+ T cell CD69 surface expression after 16 h co-incubation with APCs (n = 3–6). Data presented as mean+/− SEM; no significant differences (unpaired Student’s t test, p > 0.05) from WT controls were observed Fig. 4 figure4 CD4+ T cells deficient in cathepsin Z exhibit proficient migration in vitro; and efficient trafficking, infiltration, and reactivation in the CNS. a To determine whether deficiency of cathepsin Z affected the ability of CD4+ T cells to undergo chemotaxis in response to CXCL9, CD4+ T cells were isolated from the spleens of WT and Cat Z−/− mice and given 1 h to migrate through an ICAM coated Transwell plate in response to CXCL9 (n = 4). The negative control (−) well had no CXCL9. The positive control (+) well had no filter, allowing all CD4+ T cells to migrate through to the bottom of the Transwell. b To evaluate the ability of CD4+ T cells to infiltrate the CNS in WT and Cat Z−/− mice, MOG35–55-specific 2D2 CD4+ T cells were isolated and expanded ex vivo using IL-12 and MOG35–55 for 48 h before adoptive transfer into WT and Cat Z−/− recipient mice; alternatively, (c) to examine the capacity of Cat Z−/− CD4+ T cells to infiltrate the CNS of WT mice, WT and Cat Z−/− MOG35–55-specific 2D2 CD4+ T cells were isolated, expanded and adoptively transferred into WT recipients. b-c Six days following adoptive transfer, the 2D2 CD4+ T cells were isolated from the CNS using a discontinuous Percoll gradient, identified by flow cytometry (CD4+, Vα3.2+) and evaluated for the expression of the activation marker CD25 (n = 4). Data presented as mean+/− SEM; no significant differences (unpaired Student’s t test, p > 0.05) from the WT control were observed Mice deficient in cathepsin Z are unable to efficiently generate IL-1β during EAE and in response to inflammasome activation, and show deficiencies in Th17 polarization Although it is expected that circulating levels of proinflammatory cytokines would correspond to a decrease in neuroinflammation during EAE, it was noted that Cat Z−/− mice had dramatically lower IL-1β but comparable levels of other circulating proinflammatory cytokines during EAE (Fig. 5a). Since IL-1β has been shown to be critically important in the development of autoreactive Th17 cells implicated in the pathogenesis of MS and EAE [40,41,42,43,44,45,46], we set out to determine whether mice deficient in cathepsin Z could efficiently generate a Th17 response. When cathepsin Z-deficient mice were challenged with CFA in the absence of antigen, the draining lymph nodes contained equivalent proportions of Th1 and FoxP3+ CD4+ T cells, but proportionately reduced numbers of Th17 cells (Fig. 5b). Direct examination of a role for cathepsin Z in IL-1β production by APCs revealed significant reductions in secreted IL-1β and IL-18 following the induction of the NLRP3 inflammasome, despite equivalent expression of IL-1β mRNA in response to LPS (Fig. 5c–h). These data are consistent with a non-redundant role for cathepsin Z in the processing of IL-1β, and the IL-1β-dependent Th17 response accountable for enhanced neuroinflammation during EAE [40,41,42,43,44,45,46,47]. Fig. 5 figure5 Mice deficient in cathepsin Z have dramatically reduced circulating IL-1β during EAE and attenuated Th17 responses in vivo; consistently, APCs deficient in cathepsin Z are unable to efficiently generate IL-1β and IL-18 in vitro. a Proinflammatory cytokine levels in WT and Cat Z−/− EAE serum 15 days post induction (n = 3–6). b The percentage of Th1 (IFNγ+), Th17 (IL-17+), and FoxP3+ CD4+ T cells isolated from the inguinal lymph nodes of WT and Cat Z−/− mice 6 days after injection with CFA (n = 3). c, d Concentration of IL-1β within the supernatant of WT and Cat Z−/− (c) BMMØ, and (d) BMDC after priming with LPS and subsequent exposure to NLRP3-inflammasome activators (c) monosodium urate (MSU) or (d) ATP as quantified by ELISA (n = 5). e, f Concentration of IL-18 within the supernatant of WT and Cat Z−/− (e) BMMØ, and (f) BMDC after priming with LPS and subsequent exposure to the NLRP3-inflammasome activators (e) MSU or (f) ATP as quantified by ELISA (n = 5). g, h IL-1β mRNA levels of WT and Cat Z−/− (g) BMMØ and (h) BMDCs stimulated with LPS as determined by qPCR (n = 3–5). Data presented as mean +/− SEM; significant differences (unpaired Student’s t test, p < 0.05) from the WT control are denoted by asterisks Discussion This study provides the first experimental evidence that cathepsin Z positively contributes to neuroinflammation and the pathogenesis of EAE in a non-redundant fashion. Although phagocytic APCs express cathepsin Z at high levels, we established that cathepsin Z’s role in EAE is not mediated through antigen processing and presentation. Consistently, we showed that the enzyme’s contribution to overall proteolysis within phagolysosomes of APCs is minimal to absent—demonstrating that general protein turnover is not a primary function of this carboxypeptidase. Interestingly, we showed that cathepsin Z-deficiency leads to reduced serum IL-1β and Th17 polarization during EAE, and lowered IL-1β production by APCs in response to NLRP3-activating stimulus. Over the past decade, several pieces of evidence have emerged that implicate cathepsin Z in neuroinflammation. Cathepsin Z has been shown to be disproportionately expressed and secreted by both microglia and astrocytes in response to neuronal damage and inflammatory stimulus, both in culture and in vivo [10,11,12,13]. It was reported that dendritic cells in the aging brains of mice had increased expression of cathepsin Z that correlated with known markers of neuroinflammation [48]. Furthermore, a comprehensive comparative gene expression analysis of mouse models of MS (EAE), Alzheimer’s disease and stroke, found that cathepsin Z is one of eighteen genes whose expression is increased in all three models of neuroinflammation [38]. Whilst the expression and release of cathepsin Z has been shown to be associated with neuroinflammation, and more recently, epigenetic dysregulation of cathepsin Z with multiple sclerosis, this study provides the first experimental evidence that cathepsin Z positively contributes to neuroinflammation. Moreover, unlike typical lysosomal cysteine cathepsins, we show that cathepsin Z’s role in EAE is non-redundant, and is not mediated through antigen processing and presentation, but likely through perturbation of the IL-1β—Th17 pathway. Beyond the CNS, cathepsin Z has been associated with inflammatory conditions in other tissues. Levels of cathepsin Z (and procathepsin Z) have been shown to be significantly increased in the plasma and serum of patients who had suffered multiple traumas—which correlated with severity—and have been proposed to be used as a clinical marker for systemic inflammation [49]. Cathepsin Z has been shown to be upregulated in human gastric mucosa that is chronically infected with Helicobacter pylori, and to contribute to chronic inflammation and the development of gastric metaplasia in a mouse model of Helicobacter-induced gastritis [50, 51]. These studies suggest a broader role for cathepsin Z in inflammation, potentially mediated through a common pathway involving the generation of IL-1β. While this study identifies a mechanistic role for cathepsin Z in neuroinflammation and MS, several key questions remain unanswered. How does cathepsin Z, a strict carboxypeptidase, promote IL-1β generation, and where does cathepsin Z physically act in this pathway? In 2008, Hornung et al. reported that phagosomal destabilization and cytosolic activity of cathepsin B (or L) act to trigger inflammasome assembly in response to silica and alum, mainly based on the use of cysteine cathepsin inhibitors [52]. Subsequently, others have shown that APCs deficient in cathepsin B or L show unaltered levels of IL-1β following inflammasome activation in response to a variety of crystalline and soluble stimuli [33, 47], and that the involvement of these “typical” lysosomal cysteine cathepsins in inflammasome activation or efficiency is likely redundant. The non-redundant role of cathepsin Z in IL-1β generation suggests that this cysteine protease acts to promote IL-1β generation through a mechanism distinct from that used by cathepsin B and L within the cytosol. Alternatively, it is possible that secreted cathepsin Z (or pro-cathepsin Z) acts to enhance IL-1β generation through extracellular pathways, as has been demonstrated for cathepsin C [53]. However, since cathepsin Z is a strict carboxypeptidase and cathepsin Z-deficient APCs show reduced IL-1β release in in vitro conditions, it is unlikely that cathepsin Z directly processes extracellular IL-1β in a caspase 1-independent fashion. Another intriguing possibility is that procathepsin Z, or the cleaved proregion of cathepsin Z, enhances or triggers the assembly of the NLRP3 inflammasome through the activation of integrins on the surface of APCs in an autocrine or paracrine fashion through its evolutionarily-conserved RGD integrin binding domain [9]. Indeed, the surface protein Td92 of the periodontopathogen Treponema denticola, as well as the RGD-containing cysteine proteinase 5 of Entamoeba histolytica, have been shown to enhance inflammasome-mediated IL-1β generation through RGD-dependent activation of the α5β1 integrin on macrophages [54,55,56]. Although further insight into how cathepsin Z enhances IL-1β generation during neuroinflammation and what leads to the hypomethylation of the CTSZ locus in the human brain is warranted, the data generated in this study directly implicate cathepsin Z in the promotion of IL-1β-driven neuroinflammation. 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Download references Acknowledgements Not applicable. Funding This work was supported by the Canadian Institutes of Health Research and the Multiple Sclerosis Society of Canada. Graduate student support was provided by the EndMS training network and Alberta Innovates—Health Solutions. TR was funded by the Deutsche Forschungsgemeinschaft SFB 850 project B7 and the Excellence Initiative of the German Federal and State Governments (EXC 294). Availability of data and materials Please contact author for data requests. Authors’ contributions EA and RY conceived and designed the study. EA, RC, and RY analyzed the data and wrote the manuscript. EA, RC, BE, PT, DB, NM, CG, and AW participated in data collection. TR provided cathepsin deficient mice and contributed to the experimental design. All authors read and approved the final manuscript. Competing interests The authors declare that they have no competing interests. Consent for publication Not applicable. Ethics approval All animal research was performed in accordance with the Canadian Council for Animal Care, and protocols were approved by the University of Calgary Animal Care and Use committee. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Author information Affiliations Authors Corresponding author Correspondence to Robin M. Yates. Additional file Additional file 1: Figure S1. Mice deficient in cathepsin Z exhibit clinical signs of EAE compared to WT siblings. Although the Cat Z-/- mice used in this study were fully backcrossed to C57BL/6 (WT), to definitively rule out any anomalies resulting from background genetics or environment, EAE was induced with 50 μg MOG35-55 in CFA and 300 ng Pertussis Toxin (day 0 and 2) and scored on a standard 5 point scale. (n = 6–9). Data presented as mean+/- SEM; significant differences (Mann–Whitney U test, p < 0.05) from the WT control are denoted by asterisks (*). (PNG 140 kb) Rights and permissions Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Reprints and Permissions About this article Verify currency and authenticity via CrossMark Cite this article Allan, E.R.O., Campden, R.I., Ewanchuk, B.W. et al. A role for cathepsin Z in neuroinflammation provides mechanistic support for an epigenetic risk factor in multiple sclerosis. J Neuroinflammation 14, 103 (2017). https://doi.org/10.1186/s12974-017-0874-x Download citation Keywords • Multiple Sclerosis • Experimental Autoimmune Encephalomyelitis • NLRP3 Inflammasome • Inflammasome Activation • Experimental Autoimmune Encephalomyelitis Induction
Display Settings: Format Send to: Choose Destination We are sorry, but NCBI web applications do not support your browser and may not function properly. More information Lancet. 2000 Nov 11;356(9242):1627-31. Statins and the risk of dementia. Source Boston Collaborative Drug Surveillance Program, Boston University School of Medicine, Lexington, MA 02421, USA. Erratum in • Lancet 2001 Feb 17;357(9255):562. Abstract BACKGROUND: Dementia affects an estimated 10% of the population older than 65 years. Because vascular and lipid-related mechanisms are thought to have a role in the pathogenesis of Alzheimer's disease and vascular dementia, we did an epidemiological study of the potential effect of HMGCoA (3 hydroxy-3methylglutaryl-coenzyme A) reductase inhibitors (statins) and other lipid-lowering agents on dementia. METHODS: We used a nested case-control design with information derived from 368 practices which contribute to the UK-based General Practice Research Database. The base study population included three groups of patients age 50 years and older: all individuals who had received lipid-lowering agents (LLAs); all individuals with a clinical diagnosis of untreated hyperlipidaemia; and a randomly selected group of other individuals. From this base population, all cases with a computer-recorded clinical diagnosis of dementia were identified. Each case was matched with up to four controls derived from the base population on age, sex, practice, and index date of case. FINDINGS: The study encompassed 284 cases with dementia and 1080 controls. Among controls 13% had untreated hyperlipidaemia, 11% were prescribed statins, 7% other LLAs, and 69% had no hyperlipidaemia or LLA exposure. The relative risk estimates of dementia adjusted for age, sex, history of coronary-artery disease, hypertension, coronary-bypass surgery and cerebral ischaemia, smoking and body mass index for individuals with untreated hyperlipidaemia (odds ratio 0.72 [95% CI 0.45-1.14]), or treated with nonstatin LLAs (0.96 [0.47-1.97], was close to 1.0 and not significant compared with people who had no diagnosis of hyperlipidaemia or exposure to other lipid-lowering drugs. The adjusted relative risk for those prescribed statins was 0.29 (0.13-0.63; p=0.002). INTERPRETATION: Individuals of 50 years and older who were prescribed statins had a substantially lowered risk of developing dementia, independent of the presence or absence of untreated hyperlipidaemia, or exposure to nonstatin LLAs. The available data do not distinguish between Alzheimer's disease and other forms of dementia. PMID: 11089820 [PubMed - indexed for MEDLINE] Supplemental Content Write to the Help Desk
February 20, 2024 The McLaren F1 The McLaren F1 has a chequered history. The team was a works team for Mercedes until 2010 when they were acquired by Brawn. The Brawn team won the 2009 title with customer engines, so Whitmarsh backed out of the exclusivity deal. Afterwards, Mercedes continued to provide engines to McLaren, and sold 40 percent of its shares over a period of two years. McLaren also signed 2009 champion Jens Button to its lineup. It has been the most successful F1 since 1989, and has a reputation for being one of the world’s fastest production cars. In addition to its powerful engine, the McLaren F1 is also capable of impressive speeds. Despite its unrelentingly flashy livery, the McLaren F1 can still reach speeds of over 200 miles per hour. The car’s name, as you would expect, comes from its 6.1-litre V-12 engine. McLaren Automotive’s history has roots in the supreme talent of Bruce McLaren. The company designs cars with the driver in mind, and aims to provide the most exhilarating driving experience. Its legendary creations, such as the P1, Senna and Speedtail, have changed the landscape of the sports car industry. If you’d like to own a McLaren F1, be prepared to pay a high price for it. In the early 2000s, McLaren decided to go with a V12 engine to boost performance and reliability. The company initially sought a V12 from Honda, but they were turned down. Therefore, Murray decided to approach BMW Motorsport to develop a V-12 based on the M70. Despite the relatively high price tag, the McLaren F1 was the fastest production car ever. It won numerous championships, including Formula One, and the world’s fastest sports car. While McLaren is committed to the F1, many of its cars have undergone custom modifications and updates. While the most extreme conversions have been race cars, some of them have been rebuilt and modified to match the team’s specifications. Other cars have undergone interior and aerodynamic upgrades. These updates keep McLaren up to date with modern alternatives. This is the ultimate goal of any F1 enthusiast. With McLaren’s devoted dedication, the McLaren F1 is a must-have. The McLaren F1 has a top speed of 240 mph. Its rev limiter is set at 7500 rpm. A five-year-old prototype, the XP5, set a world record at the Ehra-Lessien test track in Wolfsburg, Germany. This incredibly powerful sports car is also affordable, with a retail price of PS460,000. In addition to being a true sports car, the McLaren F1 is a prestigious investment. The McLaren F1 has a rich racing history, which started when it was launched. The 62-unit production run ended in 1998, but there are still a large number of McLaren F1s on the road. The F1 GTR produced 28 full-on race cars, and the Sultan of Brunei owns at least eight. It is believed that around 21 of these F1s are still in the United States. About Author Leave a Reply Your email address will not be published. Required fields are marked *
0 Items Dehydroepiandrosterone What Is It Really? DHEA is a naturally occurring hormone produced in your adrenal gland. No one has yet figured out everything that it does or how important it is, but among other things, it is part of the process by which testosterone is produced. DHEA has been around a long time and has been studied for its effects on heart disease, cancer, mood, viral infections and weight, mostly in test tube and animal studies. It has been available on and off again on the underground for several years, has been used in a few small HIV studies but never in a sizeable long term study that would give us some clear answers about if and how it works, and at what dose. In 1981, The Life Extension Foundation had introduced DHEA (dehydroepiandrosterone) to its members through an article that described the multiple antiaging benefits that this hormone might produce. However, the general public did not learn about DHEA until 1996, when the benefits of DHEA were touted by the news media and in several popular books. DHEA became credible to the medical establishment when the New York Academy of Sciences published a book entitled DHEA and Aging. This book provided scientific validation for the many life extension effects of DHEA replacement therapy. The Life Extension Foundation has been investigating DHEA for more than 20 years. In more than 2,000 published studies, DHEA has been shown to have a role in improving neurological function, immune function, stress disorders, hormonal modulation and numerous diseases associated with normal aging. The most remarkable finding about DHEA comes from a human study by S.S.C. Yen and associates at the University of California, San Diego, in which 50 mg a day of DHEA over a 6-month period restored serum levels of DHEA in both men and women to youthful ranges. DHEA replacement was associated with an increase in perceived physical and psychological well-being for both men (67%) and women (84%). Increases in lean body mass and muscle strength were reported in men taking 100 mg a day, but this dose appeared to be excessive for women. DHEA (50 or 100 mg per day) was also shown to significantly elevate insulin growth factor (IGF). Aging causes a decline in IGF levels that contributes to the loss of lean body mass, as well as to excess fat accumulation, neurological impairment and age-associated immune dysfunction. Clinical studies provide evidence that DHEA improves memory, mood, and EEG readings, and may play protective role against neurodegenerative diseases. DHEA was shown to prevent pharmacologically induced amnesia and mental impairment by benzodiazepine (Valium-like) drugs. Epidemiological studies show that low DHEA levels are associated with the risk of Alzheimer’s disease, and a new study provides some molecular mechanisms for how DHEA supplementation may help in part to prevent Alzheimer’s disease. In one study, DHEA inhibited chemically induced cancers in the colon, lung, breast, and skin. When DHEA was applied directly to the skin, DHEA prevented chemically induced skin cancer. DHEA had this affect by inhibiting the binding of carcinogens to skin cells and by inhibiting the enzyme G6PDH. DHEA often declines 80-90% by age 70 or later. DHEA demonstrates a striking ability to maintain immune system synchronization. Oral supplementation with low doses of DHEA in aged animals restored immunocompetence to a reasonable level within days of administration. DHEA boosted beneficial interleukin-2 and suppressed levels of damaging interleukin-6 which is overproduced in the aged, contributing to autoimmune disease, immune dysfunction, osteoporosis and reduced healing. Suppression of interleukin-6 with 200 mg a day of DHEA was shown to be effective against systemic lupus erythematosus. Elderly people often fail to develop sufficient antibody response to vaccination. For a vaccine to work, the immune system has to generate an immune response to the vaccine. This immune response involves the production of antibodies that recognize a specific antigen on the cell of a virus or bacterium, which directs the immune system to destroy the disease-causing organism. A study in elderly volunteers showed that 100 mg a day of DHEA markedly enhanced the antibody response to the influenza vaccine. In influenza epidemics, 80-90% of mortality occurs in people over age 64. While influenza vaccines can be highly effective in young adults, 30-50% of the elderly fail to generate protective immunity. Elderly people who take an annual flu shot may want to consider taking 50 mg of DHEA daily at least two days before vaccination to help the vaccine induce an immune response. DHEA has been shown to protect against heart disease and atherosclerosis. A new study using coronary artery angiography showed that low DHEA levels are a significant risk factor for coronary artery disease. Another new study showed that DHEA inhibits abnormal blood platelet aggregation, a factor in the development of atherosclerosis, sudden heart attack and stroke. DHEA DOSING AND SAFETY PRECAUTIONS A DHEAS (dihydroepiandrosterone sulfate) blood test should be taken 3-6 weeks after beginning DHEA therapy to help determine optimal dosing. Some people neglect to test their blood levels for DHEA and wind up chronically taking the wrong dose. When having your blood tested for DHEA, blood should be drawn three to four hours after the last dose. DHEA testing may save you money if it shows that you can take less DHEA to maintain youthful DHEA serum levels. The standard blood test to evaluate DHEA status is one that measures DHEAS. The DHEAS is calculated in micrograms per deciliter (mcg/dL) of blood. The youthful ranges of DHEAS are as follows: Men 400-560 Women 350-430 People over age 40, who do not supplement with DHEA, usually have serum levels below 200, and many are below 100. Chronic DHEA deficiency is a risk factor for developing the degenerative diseases of aging according to the preponderance of evidence existing in the scientific literature. Some people obtain a baseline DHEAS blood test before beginning DHEA replacement therapy, however, based upon numerous DHEA blood tests evaluated by The Life Extension Foundation, anyone over age 40 who does not supplement DHEA is already deficient in serum DHEA. Therefore, it may be more economical to have the first DHEA blood test 3-6 weeks after initiating DHEA replacement therapy. There are precautions that should be observed that are different for men and women. Men Before initiating DHEA therapy, men should know their serum PSA (prostate specific antigen) level and have passed a digital rectal exam. Men with prostate cancer or severe benign prostate disease are advised to avoid DHEA since DHEA can be converted into testosterone (and estrogen). These sex hormones and their metabolites can promote cell proliferation. It is important to understand, however, that well-controlled studies show that serum DHEA levels are usually lower in men with malignant prostate disease compared to healthy control subjects. Therefore, men are advised to have a PSA and digital rectal exam before initiating DHEA therapy to rule out existing prostate disease, not because DHEA causes the disease. To the contrary, there is evidence indicating that maintaining youthful levels of DHEA may protect against prostate cancer. To reduce the risk that hormone modulation with DHEA could contribute to a prostate problem, men taking DHEA are also advised to take: Vitamin E 400-800 IU daily Selenium 200-600 mcg daily Mega Soy Extract 135 mcg twice daily (40% isoflavone extract) Lycopene Extract 20-40 mg daily Saw Palmetto Extract 160 mg twice daily Pygeum Extract 50 mg twice daily Nettle Extract 120 mg twice daily Gamma Tocopheral 200 mg daily Note: An aromatase inhibitor should be considered if serum estrogen levels are high. Refer to the Male Hormone Modulation Protocol for complete information about suppressing excessive estrogen levels. Men over 40 should consider checking their PSA and DHEAS serum levels every six to twelve months thereafter. Men should also periodically check their blood levels for free testosterone and estrogen to make sure that DHEA is following a youthful metabolic pathway. See the Male Hormone Modulation protocol at www.LifeExtension.com for additional hormone balance testing that can be done at the same time serum DHEA and PSA levels are being tested. Women DHEA can increase serum estrogen levels in women and eliminate the need for estrogen replacement therapy in some women. To help protect cells (especially breast cells) from excessive proliferation in response to estrogen, women taking DHEA should also take: Melatonin 500 mcg to 3 mg nightly Vitamin E Succinate 400-800 IU daily Mega Soy Extract 135 mg twice daily (40% isoflavone extract) Indole-3-carbinol 200 mg twice daily Vitamin D3 1000-1400 IU daily Women should consider estrogen and testosterone testing when they take their DHEA blood test in order to evaluate DHEA’s affect on their blood levels of estrogens. Women who have been diagnosed with an estrogen-dependent cancer should consult their physicians before beginning DHEA therapy. Some studies indicate that higher serum DHEA protects against breast cancer, but no adequate studies have been done to evaluate the effects of DHEA in breast cancer patients. If DHEA were to elevate estrogens too much, this could theoretically increase the risk of estrogen receptor positive breast cancer cells proliferating faster. Women taking DHEA should refer to the Female Hormone Modulation protocol at www.LifeExtension.com for information about restoring youthful hormone balance. Caution: DHEA can be converted into testosterone (and estrogen). These sex hormones and their metabolites can promote benign and malignant prostate cell proliferation. Men with prostate cancer or severe benign prostate hypertrophy are advised to avoid DHEA. Women with estrogen-dependent cancer should consult their physicians before beginning DHEA therapy. Individuals with existing liver disease (such as viral hepatitis or cirrhosis) might consider taking DHEA sublingually (under your tongue) or using a topical DHEA cream to reduce the amount of DHEA entering the liver. DHEA is converted by the liver into DHEA-s (dehydroepiandrosterone sulfate). Those with liver disease should carefully monitor liver enzyme levels to make sure that DHEA therapy is not making liver disease worse. This information is provided in part by http://www.lef.org/ and other various sources.

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