Immunofluorescence staining was conducted as previously described (13 (link)). Staining was conducted overnight at 4°C with anti-NeuN (1:500; ab104225; Abcam, Waltham, MA), rabbit anti-CD68 (1:500; ab125212; Abcam), rat anti-BrdU (1:500; ab6326; Abcam) and/or rabbit anti-Iba1 (1:500; ab178847; Abcam) primary antibodies. Sections were then incubated with appropriate secondary antibodies for 60 mins at room temperature (1:500; Alexa Fluor goat anti-rabbit 594, A11037; Alexa Fluor goat anti-rabbit 488, A11008; Alexa Fluor goat anti-rat 594, A11007; Invitrogen, Waltham, MA) before mounting with DAPI Fluoromount-G (0100–20; SouthernBiotech, Birmingham, AL). Fluorescent images were captured with either an EVOS fluorescent microscope (ThermoFisher Scientific, Waltham, MA) or BZ-X800E fluorescent microscope (Keyence, Itasca, IL). Image analysis was performed using FIJI (ImageJ/Fiji, version 1.53o; National Institutes of Health, Bethesda, MD).
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Immunofluorescence Staining of Neural Markers
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Corresponding Organization : Jacksonville University
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Variable analysis
independent variables
- Primary antibodies used for immunofluorescence staining (anti-NeuN, rabbit anti-CD68, rat anti-BrdU, rabbit anti-Iba1)
- Fluorescent signal intensity of stained markers (NeuN, CD68, BrdU, Iba1)
- Immunofluorescence staining protocol (as previously described in reference 13)
- Incubation time (overnight at 4°C for primary antibodies, 60 mins at room temperature for secondary antibodies)
- Mounting medium (DAPI Fluoromount-G)
- Microscopy equipment (EVOS fluorescent microscope, BZ-X800E fluorescent microscope)
- Image analysis software (FIJI/ImageJ)
- No positive or negative controls are explicitly mentioned in the provided information.
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