Immunofluorescence Assay for Neuronal Markers

Immunofluorescence (IF) assay was performed according to our previous protocol (Liu et al., 2017 (link)). The samples were first incubated with rabbit anti-BAF45D (1:100, Proteintech, Chicago, IL, United States) together with mouse anti-NEUN (1:100, Millipore, Belecula, CA, United States), mouse anti-MBP (1:500, Abcam) and mouse anti-GFAP (1:100, Proteintech, Chicago, IL, United States) overnight at 4°C. After washing in PBS, the samples were then incubated with Alexa Flour-488 anti-mouse (1:500) and Alexa Fluor-594 anti-rabbit (1:500) secondary antibodies. Nuclei were counterstained with DAPI. The IF assay for cultured cells was performed in accordance with our previous report (Liu et al., 2007 (link)) and positive staining was visualized with a NIKON Eclipse 80i fluorescence microscope and a NIKON Eclipse Ti-S inverted fluorescence microscope.