Ab186407

Western blotting was performed according to the protocol of a previous study (Wang & Wang, 2012). Cells were lysed using RIPA buffer (Solarbio) with a protease inhibitor mixture (Solarbio), and the cell lysates were then centrifuged for 30 min at 12,000 g at 4°C. The supernatants were collected. After the protein concentration was determined by BCA protein assay kit (Invitrogen), total protein (30–50 μg) was loaded for separation by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and then transferred to PVDF membranes (Millipore). Following blocking in 5% nonfat milk for 1 hr at room temperature, the membranes were incubated with a primary antibody for POLD1 (Abcam, ab186407) and β‐actin (Abcam, ab8226) overnight at 4°C. Then, the membranes were incubated with a HRP‐conjugated secondary antibody (ZSGB‐BIO) for 1 hr at room temperature after washing three times with TBST. Following washing, protein bands were visualized using a chemiluminescent substrate (Millipore) according to the manufacturer's instructions.