An 80‐day trial was carried out to evaluate the effects of replacing dietary inorganic trace minerals (ITM), with metal‐amino acid complexes (MAAC) on immune response, antioxidant capacity, and expression of cytokines genes of Nile tilapia (Oreochromis niloticus) reared in in‐pond hapas under field conditions. Quadruplicate groups of fish (93.65± 1.05 g average initial body weight ± SE) were stocked in 2‐m3 hapas fixed in a fish pond, at a density of 30 fish m‐3 (60 fish per hapa). Five isocaloric (17 MJ kg‐1), isonitrogenous (30% CP) extruded tilapia diets were produced, containing 0 (control), 25%, 50%, 75%, and 100% MAAC (Zn, Se, Cu, Fe, and Mn) as a replacement of the inorganic mineral premix. Replacement of ITM with MAAC premix enhanced the immune and antioxidant responses of Nile tilapia. The activities of lysozyme (LSZ) (96.71 U ml‐1 ± 0.69 – 120.83±1.43 U ml‐1), respiratory burst (0.54±0.08 mg ml‐1 – 3.61±0.20 mg ml‐1), alternative complement (ACH50) (34.49±56.96 % hem ml‐1– 56.96±0.79 % hem ml‐1), phagocytic cells (PA) (5.30±0.52% – 17.27±0.82%), phenoloxidase (PO) (34.96±0.96% – 54.68±0.92%), superoxide dismutase (SOD) (45.15±0.88% – 65.92±0.85%), glutathione peroxidase (GPx) (0.22±0.01 nM min‐1 mg‐1 – 1.94±0.09 nM min‐1 mg‐1), and liver function enzymes were all significantly improved (P<0.01, except for PA and GPx where P<0.05) with increasing supplemental MAAC up to 50% substitution level and decreased with further increase in MAAC to 75% and 100% levels. The expression of cytokine genes (mRNA abundance), including interferon (IFN‐γ) (2.67±0.03 – 4.62±0.08), tumor necrosis factor (TNF‐α) (2.96±0.05 – 6.73±0.05), serum alkaline phosphatase (AKP) (0.51±0.02 – 1.72±0.02), the integrated microbial genomes (IgM‐2) (22.51±0.36 ‐ 38.48±0.03) and transforming growth factor (TGF‐β) (1.84±0.02 – 7.76±0.02) genes was significantly upregulated in fish fed on MAAC–supplemented diets (P<0.05) compared with the control group. The 50% MAAC level produced the highest upregulation of these genes, whereas gene expression decreased with increasing dietary MAAC levels to 75% and 100%.