Ipsilateral basal cortical samples facing blood clots were extracted. Western blotting was performed as described previously (25 (link)). Cortical samples were homogenized and centrifuged (1,000 × g, 10 min, 4°C). The supernatant was further centrifuged, and then, the protein concentration was determined using the DC protein assay kit (Bio-Rad, Hercules, CA, USA). An equal amount of protein (50 μg) was suspended in loading buffer, denatured at 95°C for 5 min, and loaded on an SDS-PAGE gel. After being electrophoresed and transferred into polyvinylidene fluoride membranes, the membrane was blocked with nonfat dry milk buffer for 2 h and then incubated overnight at 4°C with the primary antibody for AIM2 (ab93015, 1:1,000, Abcam), pannexin-1 (ab124131, 1:1,000, Abcam), ASC (ab155970, 1:1,000, Abcam), caspase-1 (ab1872, 1:1,000, Abcam), P2X7R (1: 1,000, APR-004, Alomone, Jerusalem), IL-1β (SC-23460, 1:500; Santa Cruz), IL-18 (ab71495, 1:1,000; Abcam), and GAPDH (1:10,000, Fitzgerald, 10R-G109A). The membranes were incubated with horseradish peroxidase-conjugated secondary antibodies for 1 h at room temperature. The protein band densities were detected by x-ray film and quantified by ImageJ software (National Institutes of Health, Bethesda, MD, USA).
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