Western Blot for Bax and Bcl-2

Mitochondrial translocation of Bax and Bcl-2 was studied using Western blot. Mitochondrial pellets from each group were solubilized with radio immunoprecipitation assay (RIPA) lysis buffer containing protease inhibitors. Equal amounts of mitochondrial protein (30 μg), which was detected using a Micro BCA protein assay kit, were subjected to 10% sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and blotted into a nitrocellulose membrane. The membrane was immunoblotted for the detection of Bax and Bcl-2 protein using rabbit anti-Bax antibody (1:2,000) and anti-Bcl-2 antibody (1:2,000). The intensity of the bands in the membrane was measured using Image J.