Immunofluorescence Staining of Brain Sections

Mice were euthanized at 24 hours after surgery for IF staining, which was conducted as previously described (3 (link), 19 (link)). Brain sections with 8-μm thickness were incubated with 5% bovine serum albumin (BSA) and 0.3% Triton X-100 for 2 h at room temperature. Brain sections were incubated overnight at 4°C with primary antibodies: anti-CD68 antibody (Abcam, ab237968), anti-CD16 antibody (Invitrogen, MA1-7633), anti-CD86 antibody (Invitrogen, MA1-10299), anti-CitH3 antibody (Abcam, ab5103), anti-MPO antibody (Abcam, ab90812), anti-NE antibody (Abcam, ab68672). Sections were incubated with secondary antibodies for 2 h at room temperature. Fluoro-Jade C (FJC) staining was performed to detect neuronal damage according to the manufacturer’s protocol (Roche Inc., Basel, Switzerland). The sections were visualized using a fluorescence microscope (Leica, Germany). ImageJ software was used to analyze the results.