Comparative genomic hybridization (CGH) is a molecular cytogenetic technique that provides an overview on chromosomal imbalances within the whole tumor cell genome. This method has yet not been applied in effusion cytology. We performed CGH analysis in malignant effusions, fine needle aspirates, and imprint smears from eight ovarian adenocarcinomas, three breast carcinomas, one colon adenocarcinoma, and three malignant mesotheliomas. In part, CGH analysis from fresh frozen tissue and classic karyotyping served as controls. In this series, 14/15 cytologic specimens were suitable for extraction of high molecular weight DNA sufficient for reliable CGH analysis. CGH profiles from cytologic material were equal or even more significant in comparison with corresponding fresh frozen tumor samples. We conclude that CGH analysis from cytologic specimens may support the primary cytologic diagnosis of malignancy, especially in the differential diagnosis of benign proliferating mesothelium, malignant mesothelioma, and metastatic adenocarcinoma. CGH analysis of metastatic lesions may provide information on the site of the primary tumors and detects cytogenetic imbalances affecting oncogenes and tumor suppressor genes involved in tumor progression and metastatic spread.