Targeting Neurofilaments and Astroglia in a Sagittal Brain Section with Immunofluorescence
A sagittal section of rat brain (shown above) was immunofluorescently labeled for phosphorylated neurofilaments (expressed in neurons) with mouse anti-NF-P antibodies followed by goat anti-mouse secondary antibodies conjugated to Alexa Fluor 488. In addition, glial fibrillary acidic protein (expressed in various astroglia and neural stem cells) was targeted with rabbit anti-GFAP antibodies visualized with goat anti-rabbit secondary antibodies conjugated to Alexa Fluor 568. Cell nuclei were labeled with the popular nuclear counterstain Hoechst 33342. Images were recorded in grayscale with a 12-bit digital camera coupled to a Nikon Eclipse 80i microscope equipped with bandpass emission fluorescence filter optical blocks. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles.