Objective. The aim of this study is to explore the relationship between neuron-specific enolase (NSE) gene polymorphism and delayed encephalopathy after acute carbon monoxide poisoning (DEACMP) and provide a theoretical basis for DEACMP pathogenesis, diagnosis, and prognosis. Methods. To investigate this relationship, we screened 6 NSE single nucleotide polymorphisms (SNPs), based on the results of the previous genome-wide association studies (GWAS). A total of 1,201 patients, including 416 in the DEACMP group and 785 in the acute carbon monoxide poisoning (ACMP) group, were detected by the Sequenom MassARRAY® method. The genotype frequencies and alleles of the 6 NSE SNPs (rs2071074, rs2071417, rs2071419, rs11064464, rs11064465, and rs3213434) were compared using different genetic models. Results. In the SNPs rs2071419 and rs3213434, we found that the genotypes and allele frequencies in the two groups significantly correlated with the grouping of patients ( χ 2 = 6.596 , p = 0.037 ; χ 2 = 8.769 , p = 0.012 ). The haplotypes GGTTTC and CCTTTC of ACMP and DEACMP were different ( χ 2 = 6.563 , p = 0.010 ; χ 2 = 4.151 , p = 0.042 ). We also observed that rs2071419 and rs3213434 significantly correlated with DEACMP-increased risk in the dominant, codominant, and overdominant genetic models. In addition, we speculated that the C allele of the rs2071419 polymorphism and the T allele of the rs3213434 polymorphism in NSE may increase the DEACMP risk ( p = 0.011 , p = 0.006 ). Conclusions. The results show that rs2071419 and rs3213434 are susceptible sites of DEACMP. The NSE C allele of rs2071419 and T allele of rs3213434 and the haplotypes GGTTTC and CCTTTC may be risk factors for DEACMP.