Inducible costimulatory molecule (ICOS), a CD28 family member expressed on activated T cells, plays an important roles in T cell activation and effector function. This study was purposed to investigate the effect of blocking ICOS-B7h signal pathway by ICOS-Ig fusion protein on allogeneic reactive T cells and its mechanism. CHO cells stably and highly expressing ICOS-Ig were established, while the human ICOS-Ig fusion protein was harvested and purified from supernatant of CHO cells transfected with pSecTag2/Hygro A-ICOS-Ig. The CD4(+) cells from spleen of C57BL/6 mice were used as reactive cells, the bone marrow derived dendritic cells (DCs) from BALB/C mice were used as stimulatory cells, these cells were treated with different concentrations of ICOS-Ig or human Ig (h-Ig) as control. The results showed that the target protein with molecular weigh 54 kD and endotoxin level < 10 EU/ml was gained. The ICOS-Ig (> or = 10 microg/ml) could significantly inhibited the proliferative effect of allogeneic reactive T cells resulting from stimulation of DCs (p < 0.01). ICOS-Ig did not influence the activation of CD4(+) T cells. ICOS-Ig concentration positively related to the apoptosis of CD4(+) T cells. The percentages of CD4(+) Annexin V(+)PI(-) cells in simple stimulated group, ICOS-Ig 10 microg/ml group and ICOS-Ig 20 microg/ml group were 15.1%, 26.4% and 33.6% respectively. ICOS-Ig decreased secretion of TNFalpha and increased secretion of IL-4. It is concluded that the ICOS-Ig fusion protein has bioactivity of inhibiting T cell proliferation and altering the polarization of T helper cells to Th2 cells which promotes the apoptosis of allogeneic reactive T cells but had no effect on the activation of allo-reactive CD4(+) T cells.