Co-Immunoprecipitation is one of the classic technologies that are used widely for identification of the protein-protein interactions as well as validation. It is completely based on a particular immunological interaction that goes on between a bait protein as well as the antibody.
This process is one of the most effective as well as reliable methods that helps in detecting the interaction that is going on between the proteins. It is important that you have a sound knowledge about the complete process.
Principles of Co-Immunoprecipitation
Co-Immunoprecipitation is known to work in accordance with a number of principles. The intracellular interactions between proteins can be retained when the cells are lysed, under numerous non-denaturing conditions.
Protein X, or the bait protein, is normally captured by a particular antibody that has been stabilized to the agarose beads. If another protein is present, which is the protein Y, also known as the prey protein; they bind with each other in Vivo.
After this, this particular complex can be precipitated with the antibody. After investigating the prey protein, the interaction between both the proteins can be confirmed.
Applications of the procedure
Immunoprecipitation was primarily invented as the alternative of affinity chromatography that was used for protein production on a small scale.
The method of immunoprecipitation is one of the most popular choices for not only identifying but also separating the protein that has low abundance. The most important applications include:
- An interaction that exists between two particular proteins.
- Identifying a completely new protein with the help of a known protein.
Advantages of Immunoprecipitation
Given below is a list of the advantages that immunoprecipitation has.
- The proteins are known to interact in a physiological condition.
- The method of immunoprecipitation has a specific nature and therefore, it can be performed in a simple manner, without any hassles.
- The reagents that are normally used for this procedure can be utilized more than a single time, which makes this procedure extremely cost-effective.
- The protein complex, which is eluted, is compatible with numerous applications, which are normally used for downstream like mass spectrometry or Western blot.
- In this method, the proteins that interact with one another have been modified post-translationally and are conformationally natural.
Disadvantages of Immunoprecipitation
As important as it is to have knowledge about the advantages, it is equally important to know about the disadvantages of immunoprecipitation as well. Given below is a list of the disadvantages that you should have knowledge about.
- Weak signal from a low-affinity protein cannot be detected.
- The selection of antibody is critical and the prediction of target protein has to be appropriate.
- Antibodies that have high avidity or affinity cannot be isolated easily.
- It is not suitable for the identification of those protein interactions, which take place within a very short time.
To know more about coimmunoprecipitation, you can go through the reputed websites like Mybiosource.com. This website has information about various other tests as well.
Conclusion
It is important that you have a sound knowledge about this procedure before you decide to progress with it. Without having a proper knowledge, mistakes may be committed, which you will not know about.
