Figure 4. Colocalization of bovine RGR and cone opsins with double-label immunofluorescence. RGR and cone visual pigments were detected with DE15 (RGR), and OPN1MW/LW or OPN1SW opsin antibodies, respectively. The sections were incubated first with affinity-purified DE15 and fluorescein isothiocyanate (FITC)-conjugated anti-rabbit secondary antibody. For double-labeling, the sections were washed and incubated with the OPN1MW/LW (top panel), or OPN1SW (bottom panel) primary antibody and Alexa Fluor 568-conjugated donkey anti-goat secondary antibody. Images were obtained with a PerkinElmer 6-line spinning disk laser confocal microscope. The merged images with 4',6-diamidino-2-phenylindole (DAPI) counterstain showed intense labeling and colocalization of RGR and cone visual pigment in the outer segments of both types of bovine cone photoreceptors. Scale bar, 25 μm.