Retinal pigment epithelial (RPE) cells are responsible for helping regenerate retinal in the visual cycle, providing metabolic support to the photoreceptors, and phagocytosing the photoreceptor outer segments.¹ However, despite their important role in the retina, limited reports of RPE cell morphology exist.² RPE cells are not usually resolved in vivo in conventional scanning laser ophthalmoscopes (SLO), even those that are equipped with adaptive optics (AO), presumably because they are obscured by the waveguiding photoreceptors. Recently, in vivo techniques have been developed to image the RPE layer by using lipofuscin autofluorescence.³ Here, we combine high-resolution AO scanning laser ophthalmoscopy with simultaneous autofluorescence and reflectance imaging to resolve the human and primate RPE mosaic in vivo. Mosaics of RPE cells were imaged at different retinal eccentricities and analyzed to determine cell spacing and density. In the primate, at 10 deg from the fovea, the RPE cell spacing was 17.6 µm and the density was 2,900 cells/mm¹, while at the fovea the cell spacing was 11.6 µm and the RPE cell density was 6,300 cells/mm¹. At 15 deg from the fovea in the human eye the RPE cell spacing was 15.5 µm. This noninvasive, in vivo technique allows RPE morphology to be studied in normal and diseased retina and could be used to assess treatment efficacy in human patients or animal models of disease.