Separating adenocarcinoma of the lung from non-pulmonary adenocarcinoma or malignant mesothelioma is difficult, especially in cytology specimens. Consequently, it is important to identify markers that may facilitate this distinction. Thyroid transcription factor-1 (TTF-1) is a homeodomain containing transcription factor expressed selectively in the thyroid, lung, and diencephalon. TTF-1 is also expressed in adenocarcinomas of the lung and is widely used as a pulmonary adenocarcinoma marker in surgical specimens. However, the utility of TTF-1 has rarely been investigated in cytology. In this study, we evaluated the expression of TTF-1 in malignant pleural effusions. The primary tumors included 26 pulmonary adenocarcinomas, 26 non-pulmonary adenocarcinomas (13 breast, 5 ovarian, 2 gastric, 2 prostatic, 1 esophageal, 1 colonic, 1 pancreatic and 1 renal) and 4 malignant mesotheliomas. Immunocytochemistry was performed on sections of cell blocks, using a mouse monoclonal TTF-1 antibody (clone 8G7G3/1) and a biotin-streptavidin detection system. Nuclear immunoreactivity for TTF-1 was detected in 19 pulmonary adenocarcinomas. All non-pulmonary adenocarcinomas and malignant mesotheliomas were negative. These data indicate that TTF-1 maintains its sensitivity (73%) and specificity (100%) in cell block preparations and is useful in separating adenocarcinoma of the lung from non-pulmonary adenocarcinoma and malignant mesothelioma in cytology specimens.