We have studied the effects of low-power-density microwave (MW) radiation (continuous and pulsed wave with average power density of 10 milliwatts/sq cm and range of 5 to 50 milliwatts/sq cm; frequency, 1.0 GHz) on the uptake and action of methotrexate (MTX), the inhibition of DNA synthesis in L1210 murine leukemia cells in vitro, and the MTX treatment of mice bearing this leukemia. Using short-term tissue culture techniques, MTX concentrations of 0.2 microM, and MW exposure times of 20 min, we have found that continuous-wave low-power-density MW irradiation enhances the uptake of [3H]MTX as compared to nonirradiated controls. The enhancement is observed in only a small range of power densities (5 to 25 milliwatts/sq cm) and is in an inverted-U-shaped relationship. MW irradiation alone has an inhibitory effect on the [3H]deoxyuridine incorporation into DNA. Compared to cell suspensions treated with MTX alone, groups treated with MW irradiation followed by MTX exhibit an augmentation of inhibition of DNA synthesis as measured by [3H]deoxyuridine incorporation. Combined treatment of L1210-bearing mice with MW irradiation and MTX in vivo prolonged the duration of survival over that of animals treated with MTX alone, indicating a greater killing of leukemia cells. These results suggest that the therapeutic index of MTX may be improved by the use of MW irradiation at low power densities.