Dutch-belted rabbits with corneal epithelium either intact or debrided were injected subconjunctivally with 300 µl of one of six antifungal agents: 10 mg/ml miconazole, 5 mg/ml fluconazole, 5 mg/ml ketoconazole, 2.5 mg/ml itraconazole, and 5 mg/ml amphotericin B. At intervals of 10 min to 96 h after injection, animals were killed and corneas removed at the limbus. Three vertical strips from the right cornea and four contiguous 3-mm disks trephined from the central vertical axis of the left cornea were placed on agar plates seeded with an appropriate indicator organism. After 24 h of incubation, the zones of inhibition were measured. For itraconazole, miconazole, fluconazole, saperconazole, and ketoconazole, central corneal levels peaked by 2 h in normal and debrided corneas. Little or no drug was detectable after 4-8 h, except for itraconazole, which persisted in the cornea for at least 24 h in both normal and debrided corneas. Peak levels of amphotericin B in the central cornea were achieved after 2 h in rabbits with debrided corneas, with no drug activity measured after 8 h. There were no detectable levels of drug found in the central corneas of rabbits with intact corneal epithelium. On the basis of this pilot study, the method offers a rapid approach to the screening of antifungal agents for possible use by subconjunctival injection.