The aim of this study was to establish a method for analysis of tear film stability and thereby to investigate the effects of lubricants on corneal surface regularity and corneal epithelial damage in anesthetized rabbits with the eye held open. Saline-instilled and noninstilled eyes manifested a time-dependent increase in the corneal surface regularity index (SRI). In contrast, 0.1% hyaluronic acid (HA) prevented the increase in the SRI for up to 20 min after instillation. Methylene blue uptake into the damaged cornea was significantly greater in eyes that received saline than in those treated with HA. Whereas eyes instilled with 0.5% methylcellulose (MC) or 0.35% sodium carboxymethylcellulose (CMC) manifested an increase in the SRI similar to that apparent in those that received saline, 1.0% chondroitin sulfate (CS) prevented the increase in the SRI for up to 10 min after instillation. In conclusion, we have established a method for analysis of tear film stability in rabbits. Our results suggest that tear film stability is closely related to corneal surface damage and that topical instillation of 0.1% HA or 1.0% chondroitin sulfate warrants further investigation as a potential treatment for patients with dry eye.