Macrophage migration inhibitory factor (MIF) has been recognized as a major player in the pathogenesis of atherosclerosis. This study determined the association between polymorphisms of MIF gene and acute coronary syndrome (ACS). The polymorphism of MIF gene (rs755622, rs1007888 and rs2096525) was analyzed in 1153 healthy controls and 699 ACS cases in Chinese Han population. Plasma MIF level was also measured in part of ACS patients (139/19.9%) and healthy controls (129/11.2%) randomly. Most participants including healthy controls and ACS patients carried rs755622 GG (63.1% vs. 56.7%) and CG genotypes (33.1% vs. 38.9%) and G allele of rs755622 (79.6% vs. 76.1%, respectively), while CC genotype (3.8% vs. 4.4%) and C allele (20.4% vs. 23.9%) carriers were the lowest. Multivariate logistic regression analysis showed that carriers with rs755622 C allele had a higher risk of ACS compared to other genotypes (AOR = 1.278, 95% CI: 1.042-1.567). In addition, CC genotype carriers had the highest plasma levels of MIF than other genotype carriers. The MIF level in ACS patients with CC genotype was significantly higher than ACS patients carrying GG genotype and healthy controls carrying 3 different genotypes of MIF gene rs755622. Our findings indicate that MIF gene rs755622 variant C allele is associated with increased risk of ACS. Identification of this MIF gene polymorphism may help for predicting the risk of ACS.