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Transplantations of limbus epithelium play a steadily increasing role in the therapy of chemical burns, recurrent erosions, and impaired differentiation of the limbus epithelium (LE). To assess the vitality of LE under different culture conditions, LE was excised from 30 patients and cultivated in media with serum (F12 + 10% FCS) and without it (S4 and F12). AE5 antigen (64K keratin) was expressed by the LE specimens in these 3 media with an intensity similar to that of uncultivated specimens. All specimens strongly expressed EGF and PDGF-beta receptors under serum-free culture conditions, while serum-containing cultures reduced the expression of these receptors. Among the cells which migrated from the conjunctival preparations into the culture medium, connective tissue cells (anti-vimentin), macrophages (mac 1 and mac 2 antibodies), epithelial cells (AE5 antibody) and cells expressing class II antigen (Tü 39 and Tü 22 antibody) were determined. Only in the S4 medium were neither macrophages nor class II antigen-positive cells found. The epithelial thickness was unchanged before and after incubation with S4 medium. The two other media caused a reduction in thickness of the epithelium. The average size of the epithelial cells increased non-significantly in all cultures. Ultrastructurally, the organ cultures incubated in S4 medium showed practically no degenerative cell changes. On the basis of the criteria used here for quality checks of LE organ cultures, S4 appears to be the medium best suited on the basis of functional (PDGF-beta and EGF receptors) and morphological criteria (keratin expression, epithelial thickness and epithelial cell size).