Cd81 antibody

The serum exosomes pellet supernatant protein was loaded for electrophoresis on 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and then transferred to a 0.45-µm polyvinylidene fluoride (PVDF) membrane. The PVDF membrane was blocked with 5% non-fat milk for 1 h at room temperature and then incubated with primary antibody at 4℃ overnight. After immersed in the Horseradish Peroxidase (HRP) - conjugated secondary antibody, the target bands were detected by enhanced chemiluminescence (ECL) liquid. And the primary antibodies were purchased from abcam, including CD63 antibody (#ab217345, abcam, Cambridge, UK), and CD81 antibody (#ab79559, abcam, Cambridge, UK).

The cartilage and chondrocytes were treated with the RIPA lysis buffer (Epizyme, PC101, China) added with a protease inhibitor cocktail (Epizyme, GRF101, China) and a phosphatase inhibitor cocktail (Epizyme, GRF102). The protein contents were calculated using a BCA protein assay kit (Takara, T9300A), and the samples was instantly boiled for 10 min with the addition of loading buffer. An equal quantity of protein extracts (20 μg) was placed onto a sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) gel for electrophoresis and transmitted to a PVDF membrane. Following that, the PVDF membrane was sequentially incubated with primary and secondary antibodies. Finally, the enhanced chemiluminescence (BI, 20-500-120) was used to react with secondary antibodies and the images were acquired. The mTOR-antibody (CST, 2983), P-p70S6 (CST, 9234), Calnexin-antibody (Abcam, ab133615, USA), CD9-antibody (Abcam, ab263019), CD81-antibody (Abcam, ab109201), TSG101-antibody (Abcam, ab125011), Alix-antibody (CST, 92880), Lamp2b-antibody (Abcam, ab18529), anti-rabbit IgG, HRP-linked Antibody (CST, 7074) were used as the antibodies.