Ab72239

Total protein was isolated using RIPA lysis (Trans-Gen Biotech) according to the manufacturer’s instructions. Approximately, an equal amount of protein was separated using 12% SDS-PAGE and transferred onto PVDF membranes. After blocking with 5% non-fat milk for 1h at room temperature, the membranes were incubated with primary antibodies against CyclinE1 (ab33911, Abcam), Cleaved Caspase 3 (ab13847, Abcam), Runx2 (ab192256, Abcam), Osx (ab209484, Abcam), Ocn (ab133612, Abcam), SPTBN1 (ab72239, Abcam), TGF-β (ab215715, Abcam), p-STAT1 (ab109461, Abcam), p-SMAD3 (ab52903, Abcam), and β-actin (KM9001, Sungene Biotech, Tianjin, China). β-actin was served as the internal reference of Western Blot in this study. Subsequently, the membranes were incubated with HRP-labeled secondary antibody (1:5000) for 1 h. Finally, the detection of protein bands was performed using an enhanced chemiluminescence (ECL) kit (Santa Cruz Biotechnology, Dallas, TX, United States) according to the manufacturer’s instructions, and the bands of interest were visualized using a Bio-Rad imaging system (Bio-Rad Laboratories, Mississauga, ON, United States).

The techniques have been described previously [16 ]. The sections were incubated with pre-diluted primary Rabbit polyclonal anti-ELF antibody (ab72239, Abcam, USA) at a dilution of 1:100, with Rabbit monoclonal anti-TGF-β1 antibody (Y369, Bioworld, USA) at dilution of 1:100, at 4°C overnight. Negative controls were treated the same way, omitting the primary antibodies.