293T cells were transfected with FLAG-tagged PLK1, PLK3 or a mock vector. At 48 h post-transfection, cell lysates were subjected to IP with an anti-NPM1 antibody. The resulting immunoprecipitates were analyzed by SDS-PAGE followed by sliver staining. Gel bands corresponding to NPM1 were excised, recovered, and analyzed by LC-MS to identify serine and threonine phosphorylation sites on NPM1.