Metabolic Labeling of Protein Antigens with [³⁵S]Methionine

Metabolic labeling of antigens can be achieved by supplementation of the cell culture medium with radioactive amino acid precursors such as [³⁵S]methionine during the incubation period of target cells. In this protocol, intracellular unlabeled methionine levels are reduced by starvation of cells for 0.5-1 h before the addition of labeled [³⁵S]methionine and incubation for 0.5-4 h. Upon completion of the metabolic labeling process, cells can be prepared for immunoprecipitation by lysis or alternatively pelleted and frozen for cell lysate preparations at a later time.