The green alga Tetraspora sp. CU2551 has previously been identified and characterized as a photosynthetic microorganism with high potential for H-2 production. In the present study, cells of Tetraspora CU2551 were entrapped and immobilized in an alginate matrix with the aim to analyze the effect of cell stacking and a reduced exposure of O-2 to the cells. The results showed that the most favorable immobilization conditions were 4% (w/v) of final alginate concentration and a cell concentration of 0.125 mg cell dry wt/mL alginate with a bead diameter of 2.80-3.35 mm. The H-2 production yields increased when the immobilized cells were incubated in S-deprived medium and this could be repeated at least for 3 production times. Maximal total H-2 production reached 7.68 +/- 0.88 mL H-2/25 mL medium, corresponding to a rate of 1182.45 +/- 24.40 nmol H-2/h/mg DW. This production is about 6 times higher compared to by cells in suspension, 2-10 times higher when compared to by other green algae, and 10-50 times higher when comparing with cyanobacteria. Based on our observations, immobilized cells of Tetraspora CU2551 is considered a very promising biological system for significant photobiological H-2 production by a photosynthetic microorganism.