Glyceraldehyde 3 phosphate dehydrogenase gapdh sc 32233

Immunoblot analysis was used to investigate cell apoptotic signal proteins. RIPA buffer (Thermo Fisher Scientific) containing a protease inhibitor cocktail (Roche, Basel, Switzerland) was used to extract total proteins, and the concentration of proteins were then measured using the Bradford assay. Using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the proteins were separated and then transferred to a polyvinylidene fluoride membrane (Amersham Bioscience, Uppsala, Sweden). The membranes were incubated with primary antibody, including anti-human poly (ADP-ribose) polymerase (PARP, #9542 s; Cell Signalling, Danvers, MA), caspases-3 (#9665 s; Cell Signalling), caspase-9 (#9508 s; Cell Signalling), bax (NB100–56095; Novus Biologicals, Centennial, CO), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH; sc-32233; Santa Cruz), at 4 °C overnight. After vigorous washing with tris-buffered saline, the horseradish peroxidase-conjugated secondary antibodies, such as anti-rabbit IgG or anti-rat IgG (Invitrogen, Waltham, MA), were applied to the membranes at room temperature for 1 hrs. The specific bands were developed using the ImageQuant LAS 4000 chemiluminescence imaging equipment (GE Healthcare, Chicago, IL) and a western blot detection kit from Bio-Rad (Hercules, CA).