In a double immunofluorescence experiment, the coronal rat brain section featuring the dentate gyrus illustrated above was fixed, permeabilized, blocked with 10-percent normal goat serum, and then treated with primary chicken antibodies against NF-H (heavy chain neurofilament subunits) and mouse antibodies against calbindin (calcium-binding protein). The primary targets were then visualized with goat anti-chicken secondary antibodies conjugated to Alexa Fluor 568 and anti-mouse antibodies conjugated to Alexa Fluor 488. Cell nuclei were targeted with DRAQ5 (pseudocolored blue). Images were recorded with a 40x oil immersion objective using a zoom factor of 1.5 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.