Use of Immunomatrix Methods to Improve Protein-Protein Interaction Detection

[摘要] Immunoprecipitation (IP) and coimmunoprecipitation (co-IP) are keytechniques for studying protein-protein interactions. Thesemethods utilize immobilized protein A or protein G to isolateantibody-bound target antigens. The main disadvantage oftraditional immunoprecipitation and coimmunoprecipitation is thatthe conditions used to elute the precipitated antigen alsorelease the antibody, contaminating the antigen and destroyingthe antibody support. To overcome these problems, we describe twomethods to generate a reusable antibody support by cross-linkingthe antibody to immobilized protein A or protein G, or bycoupling it directly to the resin. Ourstudies have demonstrated that the immobilization efficiency forthe antibody coupling method was similar for several species ofantibody. Furthermore, we illustrate that using both methods ofantibody immobilization yields IP and co-IP results similar totraditional protocols but eliminate the antibody heavy and lightchains contamination.

[学科分类] 基础医学