Goat anti iba1 pab

Mice were anesthetized and perfused with phosphate-buffered saline (PBS), and the brain tissues were cut into small pieces and dissociated by enzymatic digestion using the Neural Tissue Dissociation Kit P to obtain the cell suspensions. After the digestion was completed, cells were fixed with 2% PFA for 10 min. Samples were blocked by CD16/CD32 monoclonal antibody for 10 min in FACS buffer (eBioscience, 14016182) followed by incubation with the following primary antibodies for 30 min: a) goat anti-IBA1 pAb (1:50; Abcam, ab5076); b) rabbit anti-NG2 pAb (1:50; Millipore, ab5320). The following secondary antibodies were used: a) Alexa Fluor 488 donkey antigoat IgG (1:200; Invitrogen, A-11055); b) Alexa Fluor 647 goat antirabbit IgG (1:200; Invitrogen, A27040); c) Hoechst (1:1,000; Sigma-Aldrich, 382065). The detection of EdU was performed using BeyoClick™ EdU-647 kit (Beyotime, C0081S) according to the manufacturer’s instructions.

In brief, mice were anesthetized and perfused with 4% paraformaldehyde containing 0.1 M L-lysine and 0.01 M sodium metaperiodate (PLP) as described previously (58 (link)). Immunohistochemistry was carried out according to previously published protocols (57 (link)). The following primary antibodies were used: a) rabbit anti-NG2 pAb (1:1,000; Millipore, ab5320); b) goat anti-IBA1 pAb (1:200; Abcam, ab5076); c) mouse anti-β-amyloid (1:5,000; Biolegend, 800712); d) mouse anti-p-α-syn (1:10,000 to 25,000; WAKO, 014-20281). The following secondary antibodies were used: a) Alexa Fluor 647 donkey antimouse IgG (Invitrogen A-31571); b) Alexa Fluor 647 goat antirabbit IgG (Invitrogen, A27040); c) Alexa Fluor 555 donkey antirabbit IgG (Beyotime, A0453); d) Alexa Fluor 555 donkey antigoat IgG (Invitrogen, A-31570); e) FITC-conjugated donkey antirabbit IgG (Sangon Biotech, D110051); f) Alexa Fluor 488 donkey antigoat IgG (Invitrogen, A-11055); g) DAPI (Sigma-Aldrich, D9542). Brain sections were imaged using a laser confocal microscope (Nikon TiE-A1 plus). Data were obtained and processed using ImageJ.