Fig. 3.

Immunofluorescence localization of CFH protein in human retina. Neighboring human retina sections are stained with (A) antibody to CFH or (B) antibody to CFH preabsorbed with CFH as negative control. (C) High-magnification view of the boxed area in (A). For (A), (B), and (C), left panels are the fluorescence images, with CFH labeling in green and DAPI (4′,6′-diamidino-2-phenylindole)–stained nuclei in blue; right panels are differential interference contrast (DIC) images showing the tissue morphology. In (C), the CFH signal is superimposed onto the DIC image. Labeling of CFH is intense in choroid, including blood vessels and areas bordering RPE [(A) and (C)]; this CFH signal is competed away by purified CFH protein (B), which demonstrates the labeling specificity. The fluorescence signal from RPE arises from lipofuscin autofluorescence, which cannot be competed away with CFH protein [(A) and (B)]. The black spots in DIC images correspond to melanin granules in RPE and choroids. The cell layers are indicated: GC, ganglion cells; INL, inner nuclear layer; ONL, outer nuclear layer; RPE, retinal pigment epithelium. Scale bars: 40 μm in (A) and (B), 20 μm in (C).