about immunoprecipitation - (Jan/13/2008 )
I want to know the distinguish between immunoprecipitation and co-immunoprecitation. And also the pull down assay
Hi lever! Usually you can Google immunoprecipitation and co-immunoprecipitation and get several websites with a lot of information. Also find any books available in your lab for further explanation. Here is the introduction that I wrote for my Analysis of Proteins course. I hope this helps a bit!
Immunochemical techniques are often utilized to study the interactions of antibodies with their antigens. One immunochemical method that has been performed countless times in laboratories is immunoprecipitation (IP). IP is a method by which specifically binding antibodies of peptides or proteins are precipitated out of solution using an appropriate lysis buffer. When characterizing new protein antigens, IPs are usually one of the first methods employed. In addition to characterizing protein antigens, IPs can be applied to study protein/protein interactions, to determine specific enzymatic activity, to monitor protein post-translational modifications, to determine the presence and quantity of proteins, and to enable the detection of rare proteins. To determine whether two proteins of interest are associated in vivo as well as to identify novel interacting partners of a particular protein, an analogous technique of IP is utilized – co-immunoprecipitation (Co-IP). The procedure of Co-IP is very similar to IP; however, the variance of the two techniques lies within the number of interested proteins. IP is interested in the precipitation of one protein whereas Co-IP is utilized to precipitate out multiple proteins (usually two) that interact with one another.
Generally speaking, the protocols of IP and Co-IP can be described in four simple stages: [a] lysis of the cells to release the antigens, [b] removal of nonspecific background by preclearing, [c] formation of the antibody-antigen complex and [d] purification of the immune complexes. In some cases the antigen can be labeled – most often radioactively – before lysing cells. Since antigens can be detected by methods that do not require the use of radiation, labeling of the antigen is optional.
The rest of my paper goes into detail describing the above. I have also attached two figures. Figure 1 correlates to IP and Figure 2 correlates to Co-IP. May this assist you in your future work!
I greatly appreciate your help.
Thanks very much.