The alkaline comet assay was employed to assess the pre-and post-treatment levels of in vivo DNA damage in peripheral blood leukocytes of cancer patients. During the study all patients were given antineoplastic drugs, mainly as polychemotherapy. To quantify the DNA damage, two different comet parameters were evaluated: the tail length and the tail moment. Our results indicate marked interindividual variations between baseline DNA damage in peripheral blood leukocytes recorded among cancer patients prior to the chemotherapy. After intravenous administration of various antineoplastic drugs, a significantly increased level of DNA damage in all cancer patients compared to their pre-treatment values was recorded The highest level of DNA damage was seen following administration of 5-fluorouracil, adriamycin, and cisplatin (FAP protocol). The results indicate that administration of antineoplastic drugs in standard protocols is accompanied by significant DNA damage in peripheral blood leukocytes. In order to diminish the potential risks of developing second neoplasms, a continuous biomonitoring of cancer patients after the ending of chemotherapy becomes important. Despite their limitations, present results confirm the usefulness of the alkaline comet assay as a sensitive biomarker of exposure that enables rapid and simple detection of primary DNA damage in peripheral blood leukocytes of cancer patients. Together with standard cytogenetic endpoints, the comet assay provides a powerful technique for the routine detection of critical DNA lesions produced after administration of antineoplastic drugs in the clinical settings. Teratogenesis Carcinog. Mutagen. 22:13-30, 2002.