Mouse anti gapdh

In this study, evodiamine (purity >98%), dimethylsulfoxide (DMSO), and type II collagenase were obtained from Solarbio (Beijing, China). Mouse IL-1β was purchased from Novoprotein (Suzhou, China). The following primary antibodies were utilized in the present study: mouse anti-collagen II, mouse anti-ADAMTS-5, mouse anti-COX-2, mouse anti-IκB, and mouse anti-GAPDH were purchased from Affinity Biosciences (Beijing, China). Mouse anti-iNOS, mouse anti-MMP-13, and mouse anti-Laminb1 were acquired from Proteintech (Wuhan, China). Primary antibodies to mouse anti-p65 were obtained from Cell Signaling Technology (Boston, MA, United States). Fetal bovine serum (FBS) and Dulbecco’s modified Eagle’s medium (DMEM)/F12 were acquired from Gibco (Grand Island, United States). And all ELISA kits were acquired from Cusabio (Wuhan, China).

After lysing tissue and cell samples using RIPA Lysis buffer supplemented with phosphatase inhibitors, the protein concentrations were determined using a BCA assay kit. Equal protein amounts were then separated via 12.5% SDS-PAGE, transferred onto PVDF blots, and incubated with rabbit anti-STEAP3 (#55240,1:1000, Bioss, Beijing, China) or mouse anti-GAPDH (1:3000, Affinity, Shanghai, China) overnight at 4°C. Blots were then probed for 1 h with an HRP-linked secondary antibody (1:10,000, Affinity, Shanghai, China) at room temperature, after which enhanced chemiluminescence detection of protein bands was performed. ImageJ was used for densitometric analyses. GAPDH served as a loading control.