Three human tumor cell lines (one osteosarcoma and two neuroblastoma lines) were assessed for combined interferon-beta (IFN-beta)/chemotherapeutic drug antigrowth effect under in vitro conditions. Two different methods to measure this effect were used: colony formation in soft agar and counting of cells growing as monolayers. The cells were incubated with a chemotherapeutic drug (adriamycin, dacarbazine, actinomycin D, cis-platinum, methotrexate, VP-16-213, or vincristine) at relevant concentrations for 1 h, washed twice, and incubated with IFN-beta in concentrations ranging from 100 to 1000 IU for continuous exposure. All combinations resulted in an additive or synergistic combination effect with one exception: methotrexate/IFN-beta in the monolayer method after 1 week, a combination which was additive after 3 weeks however. The combinations VP-16-213/IFN-beta and cis-platinum/IFN-beta produced the most pronounced synergistic effects. A statistical evaluation of the null hypothesis for additivity was done. These results provide a rationale for designing clinical studies combining IFN-beta with current chemotherapeutic drugs.