Antibody Labeling: The Concept And Methods
The reaction between an antibody and an antigen is a so specific process that it can always be observed under microscope for various reasons. This is made possible particularly given that the reaction take place only between an antigen and its corresponding group of antibodies. For this reason, antibody labeling is done to observe the reaction process for localization, quantification or even detection purpose.
The modern technology gives various options of antibody labeling. Some of the most common tracers used include enzymes, metallic substances like colloidal gold, radioactive biotin and even fluorochromes. An enzyme histochemical procedure under chromogenic reaction in particular works when the microscope used is a light microscope. When using fluorescent microscope however, the appropriate tracer to use id the fluorochromes. Colloidal gold works under electron miscoscope while biotin works under all the three visualization methods.
A researcher has a choice between covalent method of labeling and the non-covalent method. With covalent method, the whole process is expensive and calls for very high level of expertise. The non-covalent alternative on the other hand is much cheaper and with moderate expertise level.
In covalent method, the procedure involves the introduction of the selected labels into the antibodies via chemical processes. A group specific reagent is used to target the functional groups of antibodies so that the labeling can be done. This however involves the use of a large quantity of purified antibodies. Other than the fact that this may be expensive to obtain, purified antibodies are usually preserved with SBA.
In a covalent procedure, haptens such as biotin, enzymes, fluorophores, and DIG which are commercially available are added into a protein molecule. This is done via the process of reacting activated ester and the primary amino groups. Given that the majority of amino groups sold have BSA as a stabilizer which reduces efficiency during reaction, the LinkTM conjugation system (patented) is the technology that enables preparation of good quality conjugates even in the presence of high concentration of BSA.
The non-covalent procedure is the cheaper and more economical alternative to covalent procedure. It is now the commonly used method in most of the laboratories worldwide. In order to label primary antibodies, a fragment of a bridge monovalent Fab is used. Unconjugated primary antibodies are usually mixed with Fab fragments to obtain the labeling complex.
In both methods, localization, observation and measurement of both the antigen and the antibodies is possible with the effectiveness and efficiency highly depending on the method and substrate used. Some of the most commonly used agents when it comes to cross linking include maleimide derivatives, glutaraldehyde and m-periodate. All these are commercially available replacing many agents that were widely used in the past.
The labels to choose from are so many largely depending on the microscope to be used and the procedure to be adopted. For a success process of antibody labeling however, there are many other factors that greatly matters. These include the level of experience of the researchers, how well the laboratory is equipped, the reagents used among others. It is however important to understand that some methods allows multiple labeling of the targeted antibodies in a single incubation and sample.
The modern technology gives various options of antibody labeling. Some of the most common tracers used include enzymes, metallic substances like colloidal gold, radioactive biotin and even fluorochromes. An enzyme histochemical procedure under chromogenic reaction in particular works when the microscope used is a light microscope. When using fluorescent microscope however, the appropriate tracer to use id the fluorochromes. Colloidal gold works under electron miscoscope while biotin works under all the three visualization methods.
A researcher has a choice between covalent method of labeling and the non-covalent method. With covalent method, the whole process is expensive and calls for very high level of expertise. The non-covalent alternative on the other hand is much cheaper and with moderate expertise level.
In covalent method, the procedure involves the introduction of the selected labels into the antibodies via chemical processes. A group specific reagent is used to target the functional groups of antibodies so that the labeling can be done. This however involves the use of a large quantity of purified antibodies. Other than the fact that this may be expensive to obtain, purified antibodies are usually preserved with SBA.
In a covalent procedure, haptens such as biotin, enzymes, fluorophores, and DIG which are commercially available are added into a protein molecule. This is done via the process of reacting activated ester and the primary amino groups. Given that the majority of amino groups sold have BSA as a stabilizer which reduces efficiency during reaction, the LinkTM conjugation system (patented) is the technology that enables preparation of good quality conjugates even in the presence of high concentration of BSA.
The non-covalent procedure is the cheaper and more economical alternative to covalent procedure. It is now the commonly used method in most of the laboratories worldwide. In order to label primary antibodies, a fragment of a bridge monovalent Fab is used. Unconjugated primary antibodies are usually mixed with Fab fragments to obtain the labeling complex.
In both methods, localization, observation and measurement of both the antigen and the antibodies is possible with the effectiveness and efficiency highly depending on the method and substrate used. Some of the most commonly used agents when it comes to cross linking include maleimide derivatives, glutaraldehyde and m-periodate. All these are commercially available replacing many agents that were widely used in the past.
The labels to choose from are so many largely depending on the microscope to be used and the procedure to be adopted. For a success process of antibody labeling however, there are many other factors that greatly matters. These include the level of experience of the researchers, how well the laboratory is equipped, the reagents used among others. It is however important to understand that some methods allows multiple labeling of the targeted antibodies in a single incubation and sample.
About the Author:
If you are looking for information on antibody labeling, visit the web pages online at www.columbiabiosciences.com today. You can see details at http://www.columbiabiosciences.com now.
You are receiving this because you signed up for it on 2012-09-04 from IP 180.242.237.219
To fine-tune your selection of which articles to receive, just login here
using your username:
To unsubscribe please use the following link:
Unsubscribe
---------------------------------
New Unique Article!
Title: Antibody Labeling: The Concept And Methods
Author: Jeannie Chapman
Email: nathanwebster335@live.com
Keywords: sciene, culture, technology, education, learning, recreation, personal growth
Word Count: 535
Category: Science
---------------------------------
0 komentar:
Posting Komentar
Silahkan berikan komentar anda disini
Komentar spam akan saya hapus