The protection afforded by melatonin (MLT) against diazinon (DZN)-induced micronucleus formation, an index of DNA damage, in human blood lymphocytes was investigated. Whole blood samples were collected from five volunteers and were incubated with MLT at different concentrations (100, 200, 300, and 400 µM final concentration) for 1 h. The samples were then incubated with 750 µM DZN for 1 h. Subsequently, the lymphocytes were cultured with a mitogenic stimulant to evaluate micronucleus formation in cytokinesis-blocked binucleated cells. The incubation of lymphocytes with DZN induces additional genotoxicity. Pretreatment with MLT at these doses significantly reduced the micronucleus frequency in cultured lymphocytes (p < 0.05-p < 0.0001). The maximum decrease in the frequency of micronuclei was observed at 400 µM of MLT, which caused a reduction of 87%. MLT also exhibited an excellent and dose-dependent radical-scavenging activity against 1,1-diphenyl-2-picrylhydrazyl free radicals. Our study revealed that MLT has a potent antigenotoxic effect against DZN-induced DNA damage, which may be due to the scavenging of free radicals and increased antioxidant status. Because MLT is a natural compound and is considered safe, it can be used as a supplement to protect people exposed to chemical or environmental hazards.