Accumulation of Se in mixotrophic culture of Spirulina platensis was investigated in this study. Results indicated that glucose was better than acetate as an organic carbon source for mixotrophic culture of S. platensis. Supplementation of glucose (2 gL(-1)) significantly enhanced the biomass concentration (2.57 gL(-1)) and the production of phycocyanin (0.279 gL(-1)) and allophycocyanin (0.126 gL(-1)) in S. platensis, which were much higher than those of photoautotrophic culture (1.08 gL(-1), 0.119 gL(-1) and 0.042 gL(-1), respectively). Stepwise addition of Se during the growth phase avoided the inhibitory effect of high Se concentration on the growth of S. platensis. The Se enrichment favored the production of phycocyanin and allophycocyanin in the algal cells. The highest Se yield (1033 microgL(-1)) was obtained at an accumulative Se concentration of 250 mgL(-1), with organic Se percentage, biomass concentration, phycocyanin and allophycocyanin yields of 92.3%, 2.55 gL(-1), 0.295 gL(-1) and 0.153 gL(-1), respectively. These results indicated that the application of mixotrophic culture S. platensis with stepwise addition of Se to the medium could offer an effective and economical way for the production of high Se-enriched algal products.