Sc 33958

The Western blot was conducted as previously described.7 (link) Briefly, protein samples were obtained from the perihematomal area and BV2 cells using RIPA lysis buffer. Then, protein lysate was centrifuged for 20 min at 12,000 g at 4°C. After centrifuging, the supernatant protein solution was collected. After loading an equal amount of protein onto the gel, the electrophoresis procedure was initiated. The proteins were then transferred to PVDF membranes and blocked with a solution containing 5% bovine serum albumin (BSA) for 1 h at room temperature. After blocking, membranes were incubated with primary antibodies at 4°C overnight. The following primary antibodies were used: Dectin-1 (1:1000; ab140039, Abcam), NLRP3 (1:1000; ab263899, Abcam), ASC (1:2000; sc-33958, Santa Cruz Biotechnology), phospho-SYK (1:1000; AF3315, Affinity Biosciences), IL-18 (1:1000; 60070-1-Ig, Proteintech), GSDMD-N (1:1000; AF4012, Affinity Biosciences), caspase-1 (1:1000; 22915-1-AP, Proteintech), β-actin (1:1000; AF5003, Beyotime Biotechnology), and IL-1β (1:1000; 26048-1-AP, Proteintech). After washing three times with TBST, the membranes were incubated with secondary antibody at room temperature for 1 h and then washed a further three times. Finally, a Bio-Rad Molecular Imager was used to detect protein signals, which were quantified by Image J.