The aim of this study was to determine the in vivo healing and possible protective effects of Omega-3 against acute Tributyltin (TBT) toxicity in different tissues of rats. TBT, a tin-containing organometallic antifouling compound, is known for its adverse effects on aquatic as well as terrestrial organisms. Five groups, with seven rats each, were used in the experiments. The groups were designed as: (I) normal control, (II) vehicle control, (III) TBT (5 mg/kg/day) (IV) Omega-3 (250 mg/kg/day), and (V) TBT+Omega-3 (5 mg/kg/day + 250 mg/kg/day). Liver, kidney, spleen, and gonad tissues were assessed for their superoxide dismutase, glutathione peroxidase (GPx), glutathione reductase, and glutathione S-transferase activity levels, and changes in the glutathione (GSH) and malondialdehyde (MDA) levels, as well as serum biochemical parameters. TBT administration was found to alter the lactate dehydrogenase, total cholesterol, and low-density lipoprotein cholesterol significantly. MDA in the kidney showed a significant increase in the TBT administration compared to the control and vehicle, and a significant decrease in the Omega-3-administered groups. In the liver and spleen, the GPx showed a significantly low level of TBT compared to the control and vehicle, whereas it was significantly increased in the Omega-3 and TBT+Omega-3 groups compared to the TBT group. As a result, TBT was found to cause an increase in the lipid peroxidation, a deterioration in the lipid profile and biochemical parameters, a decrease in the GSH, and a loss of antioxidant defense enzyme activity, whereas Omega-3 was found to ameliorate some of those hazardous impacts.