Nannochloropsis sp.is commonly used as weaning feed in aquaculture, the protein content of which plays an important role in the growth and development of seedlings of aquatic animals.In this paper, three kinds of nutrients, N, P and Fe, were tested by orthogonal design methods at three levels on Nannochloropsis sp.The biomass and protein content were determined at the end of experiment.Results showed that both biomass and protein content of Nannochloropsis sp.were influenced by the levels of N, P and Fe; among the range of nutrient concentrations of this paper, P had the greatest impact on the biomass while Fe had the greatest impact on the protein content; when N, P, Fe concentrations were 24.6, 1.1 and 0.2 mg/L respectively, both the biomass yield and protein content were the highest.