The estuarine diatom Thalassiosira weissflogii (Fryxell & Hasle, 1977) has been widely used as live feed in aquaculture. The growth rate and biochemical composition of microalgae are highly influenced by environmental factors such as, light, salinity and nutrient availability. Salinity is difficult to control in some shrimp laboratories specialized in larvae production, because these laboratories depend upon the levels measured in estuaries or coastal lagoons, which are the water sources for larvae culture. The present study evaluated the effect of different salinities (25, 30, 35, 40, 45 and 50 psu), on the growth and chemical composition of T. weisflogii at three culture phases, under laboratory conditions. The highest growth rate and maximum cell density were found at 25 psu. Decrease in size and striking changes in morphology of the cells were observed at the higher salinities and drastic changes occurred at 50 psu. Protein and carbohydrate content were higher at low salinities (25 and 30 psu) during the stationary phase. The lipid production was higher at low salinities, but diminished as the phase changes occurred; in contrast, the lipid content was unaffected by the growth phase at higher salinities (≥ 35 psu). The higher growth rate and better biochemical composition were obtained at 25 and 30 psu.