An intact ciliary epithelial bilayer has been isolated from the rabbit eye by perfusion, microsurgical dissection, and recovery techniques. Vital subcellular organelles and intercellular junctions of this epithelial bilayer preparation are very well preserved. The total electrical resistance of the epithelial bilayer is 350 ohms, and the transepithelial potential is 650 microV, nonpigmented epithelium side negative. The electrical resistance is reduced by 0.2 mM EGTA and the transepithelial potential reduced by 0.1 mM ouabain. Bicarbonate depletion at a constant pH of 7.4 rapidly and significantly reduces the transepithelial potential. Carbonic anhydrase inhibitors decrease transmembrane potential by as much as 30%. These morphologic and physiologic experiments authenticate the validity of this bilayered epithelial preparation for future use in detailed studies of the mechanism of aqueous humor formation.