The current study was conducted to evaluate the effect of supplementing fish and gluten meals with chitosan and chitosan nanoparticles on the Nile tilapia growth performance, feed utilization, fatty acids profile and liver histology. The experimental design was completely randomized with a 2 x 3 factorial design; two protein sources; fish and gluten meals, and three forms of chitosan (zero-chitosan, chitosan and chitosan nanoparticles). A total number of 270 O. niloticus fingerlings were randomly distributed among 18 tanks (55 liters water capacity). The experiment lasted for 82 days. Results indicated that the growth performance parameters and protein efficiency ratio of fish fed gluten meal-based diets were improved by the addition of chitosan nanoparticles. Additionally, it was found that supplementation of gluten meal-based diet with chitosan nanoparticles increased the total omega-3 level, while decreased the omega-6/omega-3 ratio. Moreover, supplementation of gluten meal-based diets with chitosan nanoparticles improved the histological architecture of the liver and reduced fish mortality. Consequently, it is recommended to use chitosan nanoparticles as feed additives to gluten meal-based diets at a level of 0.5%.