Clinical investigations with imaging- and endoscopic techniques in order to identify the primary tumor sites in patients with CUP syndrome generally entail a significant diagnostic effort. If costs exceed <euro> 800.00, a financial loss ensues for German hospitals, as public health insurance companies do not reimburse above this amount. Combined cytological/immunocytochemical investigation of metastatic cancer cells represents a cost-effective, minimally invasive procedure to identify the probable primary cancer site that can be applied on an outpatient basis. We report on 85 fine needle aspiration biopsies of metastases to the liver, 30 to the lymph nodes and over 180 serous effusions and/or ascites with metastatic cancer cells in CUP. After conventional microscopic inspection, a routine panel of six monoclonal antibodies was applied (CK 5/6, CK 7, CK 20, Cdx 2, TTF 1 and CA 125). We were thus able to correctly identify the primary tumor sites in 90.3%, 92.0% and 85.1%, respectively, within three days. In total, 23 primary hepatocellular carcinomas could all be classified correctly, applying the antibodies HepPar 1, BerEp 4, AFP, CD 31, CD 68 and Ki 67. In addition, 141 malignant epithelial mesotheliomas were typed correctly in 97.1%, using the antibodies BerEp 4, Calretinin, Mesothelin, EMA and WT. Therefore, immunocytochemical investigation of metastatic cancer cells from fine needle aspiration biopsies or in serous effusions offers an efficient, cost-effective diagnostic alternative to imaging and endoscopic techniques in the workup of patients with CUP syndrome.