Retinal pigment epithelial (RPE) cells, retinal glia, and fibroblasts undergo marked phenotypical change when outside their usual microenvironment, as occurs in epiretinal membrane formation. To explore their phenotypic potential without the influence of other cell types, each was cultured on Gelfoam matrix and assessed immunohistochemically and ultrastructurally. All cell types demonstrated vimentin and a universal beta-tubulin epitope, TU27. RPE and retinal glial cells were positive for cytokeratin, Leu 7, and neuron-specific (gamma gamma) enolase, as were glia and fibroblasts for S-100 protein and RPE cells and fibroblasts for glutamine synthetase. RPE cells alone showed positivity for class III beta-tubulin and retinal S-antigen (monolayer cultures only); occasional retinal glia, which immunohistochemical findings suggest are Müller cell derived, demonstrated GFA protein. Therefore, class III beta-tubulin may be useful in distinguishing RPE cells from retinal glia and fibroblasts, and Leu-7 may help to identify RPE cells and fibroblasts; these cell types are difficult to distinguish in clinical material using more traditional morphological criteria.