The coronal section of rat hippocampus tissue presented above was immunofluorescently labeled for heavy chain neurofilament subunits with chicken anti-NF-H antibodies followed by goat anti-chicken secondary antibodies conjugated to Alexa Fluor 568 (yielding red fluorescence). Neurofilaments are specialized intermediate filaments that are most prevalent in the axons of neurons, but are also present in dendrites and the neuronal cell body. In addition, the calcium-binding protein calbindin was targeted in the brain section with mouse anti-calbindin antibodies visualized with goat anti-mouse secondary antibodies conjugated to Alexa Fluor 488 (green fluorescence). Cell nuclei were counterstained with DRAQ5 (pseudocolored blue), a red-absorbing fluorescent probe. Images were recorded with a 20x objective using a zoom factor of 1.7 and sequential scanning with the 488-nanometer spectral line of an argon-ion laser, the 543-nanometer line from a green helium-neon laser, and the 633-nanometer line of a red helium-neon laser. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles unless otherwise noted above.
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