Anti cyto c

For the Western blotting analysis, 80 μg of protein samples were separated on the sample buffer by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, proteins were then transferred to nitrocellulose membranes. Membranes were blocked at room temperature for 1 h in 5% non-fat dry milk in PBS, followed by incubation with anti-HK2 (1:100 dilution), anti-Cyto C (1:50 dilution, Catalog No. ab13575; Abcam), anti-voltage dependent anion channel (VDAC) (1:1000 dilution, Catalog No. ab15895; Abcam), anti-cleaved Casp 3 (1:50 dilution, Catalog No. ab2302; Abcam), and anti-β-actin (1:6000 dilution, Catalog No. a0516; Sigma-Aldrich) or anti-GAPDH (1:6000 dilution, Catalog No. G9454; Sigma-Aldrich) antibodies overnight at 4°C. After incubated with secondary antibodies, the protein bands were visualized using the LI-COR Odyssey infrared double-fluorescence imaging system (LI-COR Biosciences, Lincoln, Nebraska, USA). The relative density value of each target protein band was normalized to the density of the corresponding β-actin, VDAC, or GAPDH band.