OBJECTIVE To evaluate the effect of excimer laser photorefractive keratectomoy (PRK) on the corneal epithelial barrier function. METHODS Corneal uptake of 5,6 carboxyfluorescein (CF) was measured by Bernal and Ubels' method after excimer laser PRK in New Zealand white rabbits (N = 40). One cornea in each rabbit was treated, and the fellow cornea was used as a control. In both eyes, the central 7.0 mm of the corneal epithelium was removed. Myopic PRK treatments were performed at 37.75 microns (3.3 diopters [D]) and at 52.50 microns (5.0 D). The animals were euthanized and the eyes placed in CF for 5 minutes at 3 days, and at 1, 2, and 4 weeks following PRK. The corneas were then excised and dialyzed in balanced salt solution. The CF concentration in the dialysate was measured by fluorometry. Four corneas were also prepared for transmission electron microscopy using fixative containing ruthenium red. RESULTS Three days after PRK, CF uptake increased in all study eyes compared with normal eyes (n = 5). One week after PRK, the control corneas showed a decreased CF uptake while the study corneas still had an increased CF uptake (P < .05). Two weeks after PRK, CF uptake in corneas with a 5.0 D ablation remained increased but decreased in corneas with a 3.3 D ablation (P < .05). Four weeks after PRK, CF uptake returned to normal in all corneas. The ruthenium red penetrated into the deeper layers of the corneal epithelium 1 week after PRK; only the superficial cell layer was stained 4 weeks after PRK. CONCLUSIONS Excimer laser PRK affected the corneal epithelial barrier function at 1 and 2 weeks postoperatively even though the corneal epithelium covered the ablated area. Deeper laser ablations showed higher corneal CF uptake for longer periods than shallower ablations.