Figure 3. Western blotting and immunohistochemistry analysis of M-opsin expression treated with a protease inhibitor mixture (20 μM pepstatin A, 30 μM E64d, and 10 μM MG-132) for 24 h in Rpe65^−/− mice in retinal explants. A: Immunoblots were performed using retina-RPE choroids treated with the protease inhibitor mixture. A loading control with GAPDH was included for each immunoblot. B: Immunolocalization of M-opsin protein in the retina treated with a protease inhibitor mixture. M-opsin was stained with polyclonal anti-M-opsin antibody (red), and cell nuclei were labeled with DNA-binding dye DAPI (blue). The number of immunoreactive products increased in the cone outer segments (arrows), ONL, and OPL (arrowheads). C: Histograms showing the area of immunoreactivity of M-opsin in a 400-μm-wide section of the retina. Data are expressed as the mean±SD (n=6). *p<0.05. RPE, retinal pigment epithelium; OS, outer segment; ONL, outer nuclear layer; OPL, outer plexiform layer. Scale bars, 50 μm.