Identification and validation of protein-protein interactions by combining Co-immunoprecipitation, antigen competition, and stable isotope labeling

Abstract

Co-immunoprecipitation (coIP) in combination with mass spectrometry (MS) is a powerful tool to identify potential protein-protein interactions. However, unspecifically precipitated proteins usually result in large numbers of false-positive identifications. Here we describe a detailed protocol particularly useful in plant sciences that is based on 15 N stable isotope labeling of cells, 14 N antigen titration, and coIP/MS to distinguish true from false protein-protein interactions. © 2014 Springer Science+Business Media New York.