We performed transvitreal endoretinal biopsy in rabbit eyes to develop a reliable and safe technique to obtain retinal specimens from attached retina. Pars plana vitrectomy without lensectomy was followed by injection of Ringer's solution into the subretinal space to produce a focal retinal detachment. The apex of the focal detachment was excised by intraocular scissors and removed from the eye by pneumohydraulic expulsion. A fluid-air exchange reattached the retina. The biopsy sites were evaluated clinically and by light and electron microscopy at regular intervals up to 20 weeks postoperatively. The initial five procedures were performed without heparin in the infusion fluid, and they were complicated by severe fibrin reaction and early retinal detachment. Of the remaining 17 eyes, 15 were without intraoperative complication and maintained attached retinas. The biopsy site developed an early ring of hyperpigmentation along the border, and the biopsy bed became increasingly hyperpigmented because of cytoplasmic hyperplasia and hypertrophy of the pigment epithelium. Epiretinal membranes and subretinal neovascularization were observed histologically. Retinal biopsy specimens were reproducible and suitable for diagnostic studies.