Association of MYO7A and KLC1 with phagosomes. (A and B) Mouse primary RPE cells were fed with mouse POSs and immunolabeled with a RHO mAb (mAb1D4 in A, mAb4D2 in B; red) and a MYO7A (A) or KLC1 (B) pAb (green). (C) Proportion of POS phagosomes associated with MYO7A and KLC1 antibody labeling in different regions of cultured RPE cells. The data illustrated in the graph were from albino RPE, and the phagosomes were identified by a mixture of RHO mAb1D4 and mAb4D2. Phalloidin-TRITC was used to identify the apical region. Error bars are ± SEM; n = 10 (KLC1) or 11 (MYO7A). (D) Immunofluorescence of an RPE section from an albino mouse, double labeled with MYO7A pAb (red) and RHO mAb1D4 (green). Bracket indicates the depth of the RPE. The neural retina was detached from the RPE; the green label at the apical RPE may represent phagosomes or tips of POSs that might have remained adhered to the RPE apical surface. The green label in the choroid (Ch) represents labeling of endogenous IgG by the secondary antibody. (E) Time-lapse fluorescence microscopy of a Texas red–labeled phagosome that has KLC1-YFP associated with it. Arrows indicate direction of movement. Each frame contains red and green channel images, which were acquired sequentially, so that the red and green fluorescence appear less colocalized when the phagosome moves at a faster speed. KLC1-YFP also labels motile endosome-like organelles. Panels are from the first segment of Video 3. Bars: (A and B) 10 µm; (D) 5 µm; (E) 1 µm.