In this study, we aimed to study the possible preventive effect of docosahexaenoic acid (DHA), a dietary omega-3 fatty acid, on toxicity caused by chlorpyrifos (CPF). Six groups of Sprague Dawley rats (200-250 g) consisting of equal numbers of males and females (n = 8) were assigned to study. The rats were orally given for 5 days. The control group was administered pure olive oil, which was the vehicle for CPF. The CPF challenge groups were administered oral physiological saline, pure olive oil, or DHA (50, 100 and 400 mg/kg dosages) for 5 days. The animals were weighed on the sixth day and then administered CPF (279 mg/kg, subcutaneously). The rats were weighed again 24 h following CPF administration. The body temperatures and locomotor activities of the rats were also measured. Blood samples, brain and liver tissues were collected for biochemical, histopathological and immunohistochemical examinations. A comparison with the control group demonstrated that CPF administration increased malondialdehyde (MDA) levels in blood, brain and liver, while it reduced catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx) concentrations (p < 0.05-0.001). Advanced oxidation protein products (AOPPs) increased only in the brain (p < 0.001). DHA reduced these changes in MDA and AOPP values (p < 0.05-0.001), while it increased CAT, SOD and GPx concentrations (p < 0.05-0.001). Similarly, DHA prevented the decreases in body weight, body temperature and locomotor activities caused by CPF at 100 mg/kg and 400 mg/kg dosages (p < 0.05-0.001). Similar to the physiological and biochemical changes, the histopathological damage scores, which increased with CPF (p < 0.05-0.01), decreased at all three dosages of DHA (p < 0.05-0.01). Our findings suggest that DHA, by supporting the antioxidant mechanism, reduces toxicity caused by CPF.