Retinopathies marked by photoreceptor cell loss are the main cause of visual impairment and blindness in industrialized societies. Photoreceptor replacement strategies are examined as potential treatment options, for instance, by subretinal transplantation of photoreceptor-enriched cell suspensions, an approach which has shown beneficial effects in mouse models of retinal degeneration. As it is now possible to generate human photoreceptors through retinal organoid culture, preclinical research has shifted to human-into-mouse transplantations. Here, we describe a detailed method for human retinal organoid dissociation and fluorescent reporter-based sorting of photoreceptors. We introduce a transvitreal, micromanipulator-based transplantation technique to increase the quality and efficiency of cell injections into the mouse subretinal space, thereby increasing reproducibility and reducing experimental animal numbers. Lastly, we use a local corticosteroid to prevent immune rejection of the grafted cells in this human-into-mouse xenotransplantation setting.