Objective To investigate the effects of ISO-1,a selective macrophage migration inhibitory factor (MIF) tautomerase activity inhibitor,on the blastocysts implantation in mice and its mechanism. Methods Totally 150 pregnant mice were randomly divided into five groups. Low- dose, middle-dose and high-dose groups were given ISO-1 (2 mg/kg, 6 mg/kg, 18 mg/kg, respectively) by i.p. injection on day 3 of pregnancy and the two control groups were treated with the equal volume of 1% dimethyl sulfoxide (DMSO) or saline. A half of the mice were killed on the day 4 of pregnancy, and the uteri were excised. Structures of the endometrium were observed with HE staining. The expressions of MIF protein and mRNA were studied by the immunohistochemical staining and in situ hybridization techniques,respectively. The remaining half of the mice were killed on day 8 of pregnancy to examine the number of the embryos and calculate the uterine organ coefficient. Results Compared with the controls, there were no significant differences in the number of the embryos and the uterine organ coefficient in the low-dose group and middle-dose group( EM>P /EM>>0.05). In the high-dose group, the number of the embryos increased significantly ( EM>P/EM> <0.05), but there were no significant differences in the uterine organ coefficient compared with the controls( EM>P/EM> >0.05).The histological structure of endometrium showed no significant modifications among the groups. MIF protein and mRNA were mostly expressed in the cell aggregates scattered throughout the stroma. There were no significant differences in the average absorbance of MIF protein and mRNA in all the experimental groups compared with the controls ( EM>P /EM>>0.05). Conclusion ISO-1 may promote mice blastocysts implantation in a dose-dependent manner. ISO-1 may improve blastocysts implantation in mice by antagonizing the biological function of MIF.