Goat anti chicken af488

Immunohistochemistry was performed as previously described (Geller et al., 2019 (link)). Mice were transcardially perfused with saline followed by 4% paraformaldehyde. Brains were collected and stored in 4% PFA overnight followed by 30% sucrose for 48 h. Coronal brain sections (50 μm) were collected using a cryostat (Leica). Free-floating sections were washed in 1X PBS and incubated in Citrate Buffer (Abcam) for 15 min at room temperature (RT), followed by 2 min at 60 C° and 90 C°. Slices were incubated in blocking buffer (5% donkey serum in PBST) for 1 h at RT and incubated in primary rabbit monoclonal anti-Fgf21 (1:100, Abcam) for three nights and chicken anti-Vimentin (1:2000, Millipore) overnight at 4 C°. Sections were subsequently washed and incubated in secondary antibody donkey anti-rabbit AF594 and donkey anti-chicken AF647 (Jackson Immunoresearch) or goat anti-rabbit Texas Red and goat anti-chicken AF488 for 2 h at RT. Slices were mounted on slides with Vectashield antifade mounting media with DAPI and imaged using an Olympus BX61 Light Microscope or an Olympus FV3000 confocal laser scanning microscope. Staining for cFos was performed as previously using a cFos antibody (1:1000, Cell Signaling) (Jensen-Cody et al., 2020 (link)).