Immunohistochemistry was performed as previously published (Sun et al. 2018 (link)). In brief, 3PM imaged mice were perfused and brains were fixed with 4% PFA. Brains were sectioned coronally at 30 μm with a microtome and immunohistochemistry carried out on free-floating sections. Sections were blocked with PBS+ (10% donkey serum, 0.1% Triton-X in PBS) and incubated with primary antibodies overnight at 4 °C, followed with Alexa fluor-conjugated secondary antibodies (Invitrogen). The following primary antibodies were used: rat anti-GFAP (1:500, Invitrogen 130300), goat anti-Dcx (1:100, Santa Cruz sc-8066), goat anti-Iba1 (1:500, Abcam ab5076), rabbit anti-S100ß (Dako Z031129-2), mouse anti-Mash1 (1:100, BD Biosciences 556604). DAPI (Sigma) was used for counterstaining cell nuclei blue before mounting. Zeiss confocal scanning microscopes (Model 710 or 800) were used for image acquisition. About, 1 μm intervals were used for Z-stack scanning.
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