Donkey anti goat igg conjugated with alexa fluor 488

The injected retinal areas of the monkeys were checked by fluorescence microscopy, and the central part containing the fovea (Figure S2, shown as yellow ellipse) was dissected and fixed in 4% (wt/vol) paraformaldehyde in PBS for 2 h. Retina pieces were then dissected and re-fixed in 4% (wt/vol) paraformaldehyde for an additional 20 min. After washing in PBS, the retinas were place in 30% (wt/vol) sucrose and embedded in an embedding medium (Neg-50, Thermo Fisher Scientific). Slides containing 30-μm-thick cryosections were blocked in blocking buffer composed of 4% (wt/vol) bovine serum albumin (BSA) and 0.5% (vol/vol) Triton X-100 for 2 h in PBS. Primary antibody treatment of the cryosection slides was performed at 4°C overnight, and incubation with the corresponding secondary antibody was carried out at room temperature for 1 h. The primary and secondary antibodies used were as follows: rabbit anti-M-opsin (1:500 Millipore), rabbit anti-S-opsin (1:500 Millipore), goat anti-CNGB3 (1: 200 Santa Cruz), donkey anti-rabbit immunoglobulin G (IgG) conjugated with Alexa Fluor 594 (1:200; Jackson ImmunoResearch Laboratories), and donkey anti-goat IgG conjugated with Alexa Fluor 488 (1:200; Life Technologies.). The samples were stained with 4,6-diamidino-2-phenylindole (DAPI) and visualized with a laser-scanning confocal microscope (LSM710 Zeiss, Oberkochen, Germany).