Thrombomodulin (TM) is a glycoprotein of molecular weight 75,000 kd that is normally present in restricted numbers of cells, including endothelial and mesothelial cells. In this study, the authors tested the possibility of using anti-TM to facilitate the diagnosis of mesothelioma. All of the 31 mesotheliomas and the two mesothelioma cell lines (MS-1 and MS-2) tested were stained positively with anti-TM. The specificity of anti-TM staining in mesothelioma cells was further confirmed by in situ hybridization of MS-1 cells with a TM-specific probe. The expression of TM in MS-1 cells was increased markedly when these cells were induced by 12-0-tetradecanyl phorbol 13-acetate (TPA) to differentiate. The expression of TM in mesothelioma cells, however, did not correlate with any particular phase of the cell cycle. In an attempt to differentiate pleural mesothelioma from pulmonary adenocarcinoma, the authors compared the expression of TM, carcinoembryonic antigen (CEA), and Leu M1 in these two types of tumors. Only four of 48 (8%) pulmonary adenocarcinomas were stained positively by antibodies to TM. Therefore, immunohistochemical staining with antibodies to TM yielded 100% sensitivity and 92% specificity for diagnosis of mesothelioma. All of the mesotheliomas stained negatively for CEA and Leu M1, except for one, which showed minimal focal positivity for Leu M1. In contrast, 79% and 60% of adenocarcinomas stained positively for CEA and Leu M1, respectively. These findings suggest that immunocytochemical staining with anti-TM should be added to the battery of tests to increase the diagnostic sensitivity and specificity for differentiating mesothelioma from pulmonary adenocarcinoma.