Rabbit anti human ezh2

Cells were washed twice with ice-cold phosphate-buffered saline, and lysed in RIPA buffer (Pierce, Waltham, MA, USA). Protein lysates were separated by 10–12% SDS-PAGE, and then electrophoretically transferred to PVDF membrane (Milipore, Lake Placid, NY, USA). After blocked in 5% non-fat milk or 5% BSA, the membrane was incubated with either rabbit anti-human EZH2 (1:1000, Cell Signaling Tech., Beverly, MA, USA), rabbit anti-human EGFR (1:1000, CST), rabbit anti-human AKT and p-AKT (Ser473, 1:1000, CST), mouse anti-human XIAP (1:1000, CST), rabbit anti-human p62 (1:1000, CST), rabbit anti-human LC3B (1:1000, CST), rabbit polyclonal antibody ROCK-1 (H85, 1:200 Santa Cruz, Dallas, Texas, USA), rabbit anti-human CCND1 MET, GFAP, BMI1(1:1000, abcam, Cambridge, MA, USA), rabbit anti-human GAPDH antibody (1:1000, CST) or rabbit anti-human Tubulin (1:1000, CST), followed by HRP (horseradish peroxidase)-labeled goat-anti-mouse or goat-anti-rabbit IgG (1:5000, Santa Cruz). Protein levels were detected using ECL detection solution (Pierce) and visualized on Bio-Rad ChemiDocXRS (Bio-Rad Laboratories, Hercules, CA, USA).

Cells were washed twice with ice-cold PBS, and lysed in RIPA buffer (Pierce, Waltham, MA, USA). Protein lysates were separated by 8–10% SDS-PAGE and then were electrophoretically transferred to PVDF membrane (Millipore, Lake Placid, NY, USA). After blocked in 5% non-fat milk, the membrane was incubated with either rabbit anti-human EZH2 (1:1000, Cell Signaling Tech., Beverly, MA, USA), rabbit anti-human GAPDH antibody (1:1000, CST), followed by HRP (horseradish peroxidase)-labeled goat-anti-mouse or goat-anti-rabbit IgG (1:5000, Santa Cruz). Protein levels were detected using ECL detection solution (Pierce) and visualized on Bio-Rad ChemiDocXRS (Bio-Rad Laboratories, Hercules, CA, USA).

The procedure was described previously^{50 (link)}. Briefly, equal amounts (30-60 μg) of total protein were separated by SDS-PAGE, followed with alternative immunoblot analysis with antibodies [mouse anti-human GAPDH (Cat No. #51332, Cell Signaling Technology, Danvers, MA, USA), mouse anti-human caspase-3 and cleaved caspase-3 (Cat No. #9668, Cell Signaling Technology), rabbit anti-human EZH2 (Cat No. #5246, Cell Signaling Technology), rabbit anti-human E2F4 (Cat No. sc-1082, Santa Cruz Biotechnology, Dallas, Texas, USA), rabbit anti-human GST (Cat No. sc80998, Santa Cruz Biotechnology)]. Immunoreactive bands were then visualized, and the optical densities were measured.