Microalgae are a resource for production of renewable energy, but there are challenges that must be met to optimize the biomass production at a reasonable price. Diatoms are rich in carbon storage compounds such as carbohydrates and lipids, and studies have shown that nitrogen or phosphorus limitation may increase the cellular carbon storage. To understand better the physiological effects of resource limitation we need a controlled steady-state environment, and this can be achieved in chemostats where growth is regulated by the availability of a limiting substrate through regulation of the dilution rate. We present data from exponential fed batch-cultures of the model diatom Phaeodactylum tricornutum, to which either nitrogen or phosphorus limiting medium was added in small doses every minute over the whole cultivation period. P limitation led to a higher carbon content per cell than N limitation. A large fraction of the carbon was stored in carbohydrates in N limited cells, and this biomass may be a suitable raw material for fermentation. Independent of treatment, a lipid content of around 10% of dry weight was found in this study. In our data the biomass production expressed in terms of cell numbers was highest at intermediate dilution rates, and the chemical composition of the cells was stable over several cycles of exponential increase in culture volume and following harvests. Higher yield of storage carbon can be achieved by increasing the total biomass production instead of maximizing the per cell content of lipids or carbohydrates. (C) 2013 Elsevier Ltd. All rights reserved.