Half-life of MDM2 in RPMI 8226 MM cells. RPMI 8226 MM cells (1.0 × 10⁶ cells/mL) were cultured for 4 hours in methionine-free RPMI-1640 with L-glutamine medium supplemented with 10% dialyzed FBS, followed by culturing for 2 hours in the same medium supplemented with 50 μCi/mL ³⁵S-methionine. Cells were next cultured in RPMI 1640 with L-glutamine medium supplemented with 10% FBS, and samples of equal volume were withdrawn at 0, 10, 15, 20, 25, 30, 40, 50, and 60 minutes. Expression of labeled MDM2 was determined by immunoprecipitation of cell lysates with 1.0 μg SMP14 anti-MDM2 MoAb followed by SDS-PAGE on an 8% gel, transfer to a PVDF membrane, and autoradiography over 10 days without an image-intensifying screen. The half-life of MDM2 was 25 minutes.