Intrahepatic biliary epithelial cells were immunomagnetically purified from specimens of disaggregated human liver and were propagated in vitro. After three passes in culture, the cells were shown to be over 85% pure with contaminating leukocytes and endothelial cells constituting less than 2% of the population. Sensitive flow microfluorimetric analysis was performed after immunofluorescence labeling to quantify expression of both class 1 and class II major histocompatibility (MHC) antigens and the adhesion molecules intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and lymphocyte function-associated antigen-3 (LFA-3). It was shown that resting human intrahepatic biliary epithelial cells (HIBEC) expressed class 1 MHC antigens, ICAM-1, and relatively low levels of LFA-3. Stimulation with tumour necrosis factor-alpha (TNF-alpha) upregulated expression of ICAM-1, whereas Interferon-gamma (IFN-gamma) and a combination of IFN-gamma and TNF-alpha upregulated class I MHC antigens and ICAM-1 and induced class II MHC molecules. The level of expression of MHC antigens and of ICAM-1 and LFA-3 after stimulation of HIBEC with combined IFN-gamma and TNF-alpha was comparable with the expression of these antigens by an Epstein-Barr virus-transformed B-cell line. The phenotypic similarity between cytokine-stimulated HIBEC and a known antigen presenting cell is consistent with a potential role for biliary epithelial cells in antigen presentation.