Co-Immunoprecipitation Protocol

Co-immunoprecipitation protocol (Co-IP) is a widely used procedure to analyze protein interaction. The procedure is generally comprised of three steps. An antibody is introduced to a cell lysate, the antibody-protein complex is chemically bound and the protein is identified through analysis.

Co-Immunoprecipitation Summary

• Co-IP is comprised of a series of steps to ensure accurate test results. The first step requires an antibody specific to protein to be added. In the second step, protein-G or protein-A sepharose is used to bind the antibody-protein complex. Finally, sequencing a purified protein band, or the Western blot analysis, is used to identify the co-precipitated protein.

General Steps

• The protocol can be divided into three general steps in term of the mechanics of the process. The first step is work flow to complete the procedure in the least amount of time. The second step is general considerations for the procedure. The final step is troubleshooting.

Specific Steps

• The work flow begins with preparing the culture for the blotting analysis and concludes with determination of the presence of any proteins. General considerations consist of key steps such as pre-cleaning, the choice of buffers and selection of Protein A or Protein G Resin. Troubleshooting deals with issues such no detection of protein, binding of the antibodies to proteins, antibody fragments and the possibility of an unsuccessful Co-IP.