Clinical trials using interferon to treat human malignancies are currently hampered by limited supplies of the compound. We have utilized a human tumor cloning system as an assay for the antitumor effects of human leukocyte interferon. Interferon was tested against 62 patients' tumors growing in this soft agar system. A dose-dependent cytotoxic effect of interferon was noted against only five of the patients' tumors. A greater than or equal to 70% decrease in tumor colony-forming units (TCFUs) was noted with one lymphosarcoma cell leukemia, one small cell lung cancer, one adenocarcinoma of the lung, one breast cancer, and a pancreatic cancer. One patient had his tumor cultured in vitro and had a clinical trial with interferon. This patient whose tumor demonstrated in vitro sensitivity had a clinical antitumor effect with interferon therapy. The in vitro results in this study suggest that the human leukocyte interferon currently available has a low level of activity in a human tumor cloning system. Additional testing is needed to determine whether the cloning system can identify the patient(s) who might have an antitumor effect from the interferon.