Sc 7177

Osteoblast cells were lysed with 100 µL/50 mL protein extract lysates. Briefly, after optical density (OD) was measured by a microplate reader (Varioskan LUX, Thermo Fisher Scientific) at 490 nm, proteins were separated by sodium dodecyl sulfatepolyacrylamide gel electrophoresis on 10% polyacrylamide gels and transferred to polyvinylidene fluoride membranes. After being blocked with 5% skim milk/BSA, the membranes were incubated with primary antibodies for EZH2 (1:1000, sc-131324, Santa Cruz Biotechnology, Delaware Ave Santa Cruz, CA, USA), RUNX2 (1:1000, sc-7177, Santa Cruz Biotechnology), OSTERIX (1:1000, cs-9272, Cell Signaling Technology, Inc., Boston, MA, USA), and glyceraldehyde-3-phosphate dehydrogenase (GADPH, 1:1000, sc-131325, Santa Cruz Biotechnology) overnight at 4℃. Then membranes were washed with Tris Buffered Saline Tween (TBST) and incubated with horseradish peroxidase-conjugated secondary antibody (1:5000, Shanghai Solar Biological Reagent Co., LTD., Shanghai, China). Protein bands were visualized using a gel imaging system (E-Gel Imager, Thermo Fisher Scientific).