Interleukin-12 (IL-12) has been evaluated in both preclinical and clinical immunotherapy protocols as a potential therapy for melanoma. However, delivery of IL-12 in the form of recombinant protein can result in severe toxicity, and gene therapy has had limited success against B16.F10 murine melanoma. This study investigated the therapeutic effect of delivering a plasmid encoding IL-12 followed by electroporation on primary and secondary tumors. Three treatments of intratumoral (i.t.) plasmid injection and electroporation resulted in 80% of mice with B16.F10 melanoma tumors being tumor free for >100 days (cure). The "cured animals" were resistant to challenge with B16 cells. In a separate experiment, B16 cells were injected on the opposite flank of the treated tumor on the day of treatment. Eighty-seven percent of control mice developed a distant tumor while only 43.8% of mice receiving two or three i.t. electroporation treatments developed a distant tumor. For examination of tumor development in the lungs, mice were injected intravenously with B16.F10 cells then treated with i.m. injections of plasmid with or without electroporation. Only 37.5% of mice receiving i.m. injections and electroporation developed nodules in the lungs compared to 87.5% of mice in the no-treatment group. The results show that administration of a plasmid encoding IL-12 with electroporation has a therapeutic effect on primary tumors as well as distant tumors and metastases.