β actin polyclonal antibody

Cells were harvested and lysed in PRO-PREP protein extraction solution (iNtRON Biotechnology). Cell lysates were scraped, and protein concentrations were measured with the Bradford assay. Equal amounts of protein were denatured in 5×protein sample buffer (ELPIS biotech, Daejeon, Korea), separated by SDS-PAGE on 12% polyacrylamide gel, and transferred to Immun-Blot®PVDF membranes (Bio-Rad, Quarry Bay, Hong Kong). The membranes were blocked with 3% bovine serum albumin, and probed with the following primary antibodies overnight at 4°C: anti-leptin antibody (Abcam), anti-p-JAK2 antibody (Abcam), anti-p-STAT3 antibody (Cell signaling, Danvers, Massachusetts, USA), anti-Notch1 antibody (bioworld, Dublin, Ohio, USA), anti-NICD antibody (Abcam), anti-Jagged1 antibody (Abcam), anti-survivin antibody (Enzo Life Sciences, Farmingdale, New York, USA) or β-actin polyclonal antibody (Abcam). After washing, the membranes were incubated with goat anti-rabbit IgG antibody (H+L), HRP conjugated (Bioss Antibodies, Woburn, Massachusetts, USA) for 1 hr at room temperature, blots were visualized using Chemiluminescent Sensitive Plus HRP Microwell and/or Membrane Substrate (SurModics, Eden Prairie, Minnesota, USA) and signals were measured with a Chemi-Doc® (Bio-Rad, Hercules, California, USA) [26 (link)].