Primary cultures are introduced as a method for an immunocytochemical and functional characterization of epithelial cells (ECs) from human thymic epithelial tumors. Neoplastic ECs were obtained after enzymatic digestion of the tumor tissue with dispase. The ECs were kept in culture for up to 1.5 months. Over this period a progressive decline in their proliferation rate was observed. In all five cases studied, ECs showed a co-expression of keratin and vimentin intermediate filaments in vitro as well as strong expression of major histocompatibility complex (MHC)-class I antigens and a progressive loss of MHC-class II antigens. Acetylcholine receptor (AchR)-epitopes were detected immunohistochemically by using monoclonal antibodies (mAbs) to the cytoplasmic site of the alpha-chain if the AchR. Epitopes were found in three of five thymomas in vivo and to a varying degree in all five cases in vitro.