Anti rabbit ask1

Brain tissues were fixed with 4% paraformaldehyde for 24 h and then incubated with 20% and 30% sucrose solutions for 24 h. Next, the brains were sectioned into 25 μm sections, and the sections were washed 3 times with a phosphate-buffered saline (PBS) solution. After washing, the sections were permeabilized with 1% Triton X-100 and blocked with donkey serum for 90 min, followed by incubation with primary antibodies overnight at 4 °C. The primary antibodies used included: rabbit anti-ASK1 (1:100, Abcam, US), goat anti-Iba1 (1:1000, Novus, US), mouse anti-NeuN (1:2000, Abcam, US), rat anti-CD16 (1:100, BD, US), rabbit anti-CD206 (1:200, Abcam, US), rabbit anti-pJNK (1:200, Affinity, China), rabbit anti-pp38 (1:200, Affinity, China), rabbit anti p–c jun(1:200, Cell Signaling Technology, US) and rat anti-CD68(1:500, Bio-Rad, US). The next day, the sections were incubated with secondary antibodies in the dark for 60 min at 37 °C. The following secondary antibodies were used: Alexa Fluor 594-labelled goat anti-rabbit IgG (Proteintech; 1:1000), fluorescein isothiocyanate (FITC)-conjugated goat anti-mouse IgG (Proteintech; 1:1000), and Alexa Fluor 647-labelled goat anti-mouse IgG (Proteintech; 1:1000). Finally, the immunofluorescently labelled sections were captured using a laser scanning confocal microscope (Nikon, Germany).