Labeling cultured cells with 32Pi and preparing cell lysates for immunoprecipitation

Signal transduction pathways may involve protein phosphorylation at one or more residues. Detection of phosphorylation involves labeling with radioactive inorganic phosphate and subsequent immunoprecipitation with appropriate antibodies. This unit describes labeling conditions for adherent and nonadherent cells and preparation of the lysates from these cells for immunoprecipitation. The labeling method is appropriate for labeling other phosphorylated component of cells, but alternate methods are necessary for their detection.