An experiment was conducted for in-situ rearing of GIFT tilapia to evaluate the efficacy of bioflocculating agent chitosan at various dosages in the inoculum. Growth and physiological responses of GIFT Tilapia was studied in a biofloc system developed with a bioflocculating agent based inoculum. A dose optimization study was carried out to determine the optimum dose of chitosan in the inoculum using 3 different doses viz. 30, 60 and 90 ppm. Based on the results of dose optimization, the second experiment was conducted using 10, 20 and 30 ppm of chitosan dosage in the inoculum designated as T1, T2, T3, respectively, and one control (without chitosan) with in-situ rearing of tilapia. The tanks (12 Nos) were stocked randomly with an equal number of fishes of initial mean weight (1.70 ± 0.36 g) at 50 numbers per 250 L of water and C: N ratio was maintained at 15:1 using tapioca flour as carbon source. The biofloc developed with the inoculum using 30 ppm (T3) chitosan showed highest floc volume, highest flocculation activity (83.82 ± 1.84%) and highest total suspended solids level (550.67 ± 32.23 mg L - 1). Higher body weight gain (11 g), specific growth rate, feed conversion ratio (1.5) and feed efficiency ratio were observed to be significantly similar for 20 and 30 ppm chitosan supplemented biofloc groups, where as they indicated higher response compared to 10 ppm (T1) and 0 ppm (C) chitosan supplemented group. The activities of amylase was increased in T2 and T3 groups, while the activities of hepatic and gill superoxide dismutase (SOD) and lactate dehydrogenase (LDH) were significantly lower in T1, T2 and T3 groups compared to control. However, activities of protease, lipase and catalase, serum cortisol and glucose were non-significant. The study concluded that the introduction of bioflocculating agent chitosan at a dosage of 20–30 ppm in the inoculum for in-situ biofloc technology can enhance growth, feed utilization, amylase activity and ameliorating stress in GIFT tilapia.