Background: This study was performed to evaluate any synergetic effects of mitoxantrone (MX) and gold nanoparticles (GNPs) as dual therapeutic approach, along with microwave (MW) hyperthermia for melanoma cancer. Methods: Various tests were performed on the DFW melanoma cell line in the presence of MX and different concentrations of GNPs, with and without MW irradiation. MTT [3-(4,5-dimethylthiazol–2-yl)-2,5-iphenyltetrazolium bromide] assays were conducted to evaluate the effectiveness of the used therapeutic methods in terms of cell survival. Relative lethal synergism (RLS) was calculated as the ratio of cell death following hyperthermia in the presence of a treatment agent to that after applying hyperthermia in the absence of the same treatment agent. Results: Results showed MX and GNPs under MW irradiation to provide maximum cell death (P < 0.001 compared to the other groups). The mean RLS for MW hyperthermia along with the MX-GNP combination was 4.14, whereas in the absence of GNP the value for MX chemotherapy was 0.94. Conclusion: MX chemotherapy in the presence of different concentrations of GNP did not alter cell survival as compared to in its absence.