Mouse anti pgp9

Skin was dissected from 8 week old adult mice and post-fixed in 4% PFA for 30 min at RT. DRG were cryo-protected overnight at 4°C in 30% sucrose-PBS, embedded in OCT, and then sectioned at 18 μm onto slides. Briefly, slides were blocked in 5% normal goat serum in PBST (0.1% Triton X-100) and incubated overnight at 4°C in 1:1000 primary antibody in blocking buffer. Slides were washed 3X in PBS then incubated 45 min at RT in 1:1000 secondary antibody. Slides were washed 5X in PBS and mounted in Fluoromount-G +DAPI with No. 1.5 coverglass. Primary antibodies used: Rabbit anti-DsRed (Clontech #632496), Chicken anti-NefH (Abcam #4680), Chicken anti-β-tubulin III (Abcam #107216), mouse anti-PGP9.5 (Abcam #8189). Secondary antibodies used: Goat anti-Mouse Alexa 488 (Abcam #150117), Goat anti-Rabbit Alexa 594 (Invitrogen #R37117), Goat anti-Chicken Alexa 488 (ThermoFisher #A11039). For co-localization analysis, only fibers for which >50% of the length of the visible fiber contained co-localized (white) pixels were counted. Image analysis was performed using FIJI.