Ab0054

RIPA buffer (Solarbio, Beijing, China) was utilized to separate total proteins from cell lysates. After resolution using 8% sodium dodecyl sulfate–polyacrylamide gel electrophoresis, proteins were shifted onto 0.22 μm PVDF membranes (Millipore). Following 1 h of blocking in 1× TBST buffer containing 5% nonfat dry milk, an overnight incubation of the membranes was performed at 4°C using primary antibodies to NF-κB p65 (#8242, 1:1000, CST, USA), IκBα (#4814, 1:1000, CST, USA), p-IκBα (#2859 1:1000, CST, USA), IKKβ (#8943 1:1000, CST, USA), p-IKKα/β (#2697 1:1000, CST, USA), NF-κB p50 (CY5040, 1:2000, Abways, China), RAB10 (ab237703, 1:1000, Abcam, USA), E2F2 (ab138515, 1:1000, Abcam, USA), Lamin B1 (AB0054, 1:1000, Abways, China) and β-actin (AB0035, 1:1000, Abways, China). After a triple ten-minute rinse using 1× TBST buffer, the membranes were proceeded 1 hour incubation at 37°C protected from light utilizing the corresponding DyLight 800-labeled goat anti-mouse or anti-rabbit antibodies (1:7000, abbkine, China). Finally, the blot image was recorded with the Odyssey Infrared Laser Scanner (LICOR Biosciences). Three replicates were required for each experiment. β-Actin levels were employed to normalize protein levels of the whole cell lysis and Lamin B1 for nuclear extract.