AF2 antibody shows detection of endogenous TRAF2 in whole HeLa cell lysates. Lane 2 shows endogenous TRAF2 detected with antibody at 47 kDa (arrowhead). Lane 4 shows no reactivity when blot is incubated with immunizing peptide. The identity of lower molecular weight band in lane 2 is unknown. Briefly, each lane contains approximately 14 µg of lysate. Membranes were blocked in 3% BSA-TBS 30 min. at room temperature. Primary antibody was used at a 1:500 dilution in 3% BSA-TBS and reacted overnight at 4°C. The membrane was washed and reacted with a 1:20,000 dilution conjugated Gt-a-Rabbit DyLight 649 for 1 hr at room temperature. Molecular weight estimation was made by comparison to prestained MW markers in lanes 1 and 3. Fluorescence image was captured using the VersaDoc Imaging System developed by Bio-Rad.