The study aimed to investigate the effect of fish oil (FO) replacement with soybean oil (SO)on growth performance, biochemical and hematological parameters, and immune response of Nile tilapia. Ninety fish, weighed 20.1±0.2 g, was assigned into 3 groups (3 replicates each). Three dietary treatments contained FO (5%), SO (5%) or an equal mix(2.5+2.5%) of fish and soybean oils (FSO). The fish was fed on iso-caloric (3147 Kcal/kg) and iso-nitrogenous (30.62% crude protein) diets. The experiment lasted for 8 weeks. The average daily gain of fish fed FO was the greatest (0.61±0.03g) followed by FSO (0.56±0.01 g) then SO (0.54±0.01 g). Also, FO significantly improved feed conversion ratio (1.66±0.04) as compared with SO (1.82±0.02) or FSO (1.78±0.07). FO significantly stimulated feed intake as compared to SO. FO significantly increased ash and decreased fat contents in the carcass. A significant increase in the number of red blood cells, packed cell volume and hemoglobin concentration was achieved in the group fed FO than those fed on SO or FSO. However, SO increased heterophils and decreased lymphocytes as compared with FO and FSO. Also, serum protein was significantly increased in fish fed FO than the other two groups. FO significantly decreased serum triglycerides, cholesterol, and creatinine than the other two groups. Moreover, FO significantly increased serum antibacterial activity (48.5%) than SO (44.4%) or FSO (45.8%). It could be concluded that the complete replacement of FO with SO decreased growth performance and feed utilization of the Nile tilapia. However, FSO could be practically utilized in fish diets without a harmful effect on the growth performance of Nile tilapia fish.