The appearance of cell surface glycoconjugates (detected by fluorescein-isothiocyanate-conjugated lectins and bullous pemphigoid antibody) was serially examined in mouse epidermal cell cultures treated with trans-retinoic acid and aromatic retinoic acid (etretinate) and in cultures maintained under low calcium conditions. The changes in lectin staining occurred in concert with the process of differentiation as assessed by cell morphology and colony growth characteristics, and they correlated with the patterns observed in whole mouse skin. The keratocyte cultures treated with retinoic acid showed delayed and reduced differentiation and stratification, and this was associated with markedly reduced binding of lectins specific for N-acetyl-glucosamine and fucose. The low calcium concentration produced similar changes. Thus, the loss of surface glycoconjugates in the epidermal cell culture system was not specific for either retinoic acid or low calcium, but correlated with the degree of cell differentiation.