Anti gapdh antibody

Extraction of the total protein from cells in RIPA lysis buffer (Beyotime, Shanghai, China) containing the cocktail. Approximately 25 μg of protein was detached via a 10% sodium dodecyl sulfate-polyacrylamide electrophoresis (SDS-PAGE) gel, and further transmitted to 0.45 µm PVDF membranes (Millipore, MA, USA) and blocked in TBST containing 5% fat-free milk for about 1 hour. Later, incubate the membranes with antibodies. Visualization of immunological assays was performed by using a chemiluminescent ECL reagent (Beyotime, Shanghai, China). Primary antibodies were used as follows: anti-AGO2 antibody (Abcam, Cambridge, UK), anti-IFIT2 antibody (Proteintech, Wuhan, China), anti-RUNX2 antibody (Abcam, Cambridge, UK), anti-ALP antibody (Abcam, Cambridge, UK), anti-OPN antibody (Abcam, Cambridge, UK), and anti-GAPDH antibody (ZenBioscience, Chengdu, China).

Cells were lysed, denatured, separated and transferred. The following three primary antibodies were used: anti‐SATB2 antibody (1:1000; Abcam, Cambridge, UK), anti‐BMP9 antibody (1:1000; ThermoFisher Scientific, Waltham, USA) and anti‐GAPDH antibody (1:3000; Zen Bioscience, Chengdu, China). Chemiluminescence (Beyotime, Shanghai, China) reagent was used to visualize the presence of the proteins of interest. Image J software was employed for protein band quantification.