Te 2000

Mice were euthanized 24–28 hours after Sindbis virus injection by brief isofluorane anesthesia followed by decapitation, and the brains were immediately extracted. The olfactory bulb was cut off and either fixed in 4% paraformaldehyde (PFA, Electron Microscopy Sciences) in PBS, post-fixed for 24 hours, and cryoprotected before frozen, or directly frozen as previously described (Chen et al., 2019 (link)). The bulb hemispheres were then sliced into 20 µm thick sections and processed for BARseq. BARseq library preparation and sequencing was done as previously described (Chen et al., 2019 (link)). In situ sequencing was performed with a custom-made automated fluidic sequencing system on either a Nikon TE-2000 or an Olympus IX-81 with a Crest X-light v2 spinning disk confocal microscope using a LDI 7-laser light source. Filters and lasers used for each imaging channel documented in Table S4.

Inverted fluorescent microscope (Nikon TE2000) and confocal microscope (Olympus Fluoview) were used for cell imaging. 3T3 Swiss mice fibroblasts were seeded at 50,000 cells in 1000 μL of media per dish in DMEM complete media and incubated overnight in 35mm petri dishes with No. 1.5 coverslip as a bottom (MatTek Corporation). The following day cells were incubated with studied peptides (PBS, 0.1 μM to 100 μM) for up to 30 min. After incubation, cells were washed three times with PBS and 1 ml of PBS was added to the plate. The cells were imaged immediately with an Olympus Fluview confocal microscope at 20X, 40X and 100X oil immersion objectives. If DAPI (Molecular Probes) staining was performed, the cells were fixed with glutaraldehyde (Aldrich), washed with PBS and stained.