Rabbit anti trpv4

The antibodies used were: rabbit anti-TRPV4 (#ab39260, Abcam, Cambridge, Britain); rabbit anti-iNOS (#13120, Cell Signaling Technology, Danvers, MA, USA); mouse anti-F4/80 (#sc-377009, Santa Cruz, Santa Cruz, CA, USA); rabbit anti-COX2 (#ab15191, Abcam, Cambridge, Britain); rabbit anti-GAPDH (#8884, Cell Signaling Technology, Danvers, MA, USA); rabbit anti-NLRP3 (#13158, Cell Signaling Technology, Danvers, MA, USA); rabbit anti-CASPASE-1 (#22915-1-AP, Proteintech, Wuhan, China); and rabbit anti-ASC (#10500-1-AP, Proteintech, Wuhan, China).

Brain slices were prepared as previously described,
²¹ (link) and 0.3% Triton ×100 was used to penetrate 15 min. Brain slices (30 μm) were blocked with QuickBlock™ (Beyotime, China) for 1 h, and then incubated in a refrigerator with primary antibodies at 4°C overnight. We performed single immunofluorescence staining using rabbit anti‐Iba1 (1:1000, Abcam, USA). Performed double immunofluorescence staining using mouse anti‐Iba1 (1:100, Santa Cruz Biotechnology, USA) with rabbit anti‐TRPV4 (1:200, Abcam, USA), rabbit anti‐Iba1 (1:1000, Abcam, USA) with mouse anti‐CD86 (1:50, Santa Cruz Biotechnology, USA), and rabbit anti‐Iba1 (1:1000, Abcam, USA) with goat anti‐CD206 (1:250, R&D Systems, USA). After washing the brain slices with PBS containing 0.3% Triton ×100, Alexa 594‐conjugated goat anti‐rabbit antibodies (1:1000, Life Technologies Corporation, USA), Alexa 555‐conjugated donkey anti‐goat antibodies (1:1000, Life Technologies Corporation, USA), and Alexa 488‐conjugated goat anti‐rabbit antibodies (1:1000, Life Technologies Corporation, USA) were added and incubated at room temperature for 2 h. The nucleus was stained with DAPI. Using laser scanning confocal microscopy (Leica SP‐8) to take pictures. ImageJ was used for analysis.