This study was aimed to investigate whether polymorphisms in HLA DRB1 genes and TNFα gene promoter ( 238 and -308), and their combined haplotypes, could be risk factors for neural tube defects (NTD) in humans. The study included 111 children with NTD, the control group consisted of 113 healthy inhabitants of the Southern Poland. TNFα gene promoter polymorphisms were detected with RFLP, and HLA DRB1 were genotyped with PCR sequence-specific oligonucleotides reverse dot-blot kits. Statistical analysis involved descriptive statistics, exact Hardy-Weinberg test, Fisher’s exact test (with Bonferroni correction if applicable), χ2 (with Yates’ correction if applicable), Cochrane-Armitage trend test, G-test (log-likelihood ratio test) and permutation test. Haplotypes were calculated with PHASE v. 2.1. software. No differences were identified between the controls and NTD children in frequency of individual HLA DRB1 alleles, their global distribution, as well as frequency of HLA DRB1 homozygotes and heterozygotes. No difference was found between the controls and NTD patients for TNFα gene promoter polymorphisms. A haplotype DRB1*07-TNFα( 238)G- TNFα(-308)G was found more frequently in the whole NTD group vs. controls. NTD patients and controls did not differ in terms of haplotype distribution. Conclusions: no association of HLA DRB1 and TNFα gene polymorphisms was demonstrated with NTD in humans. No conclusions can be drawn about association of a haplotype DRB1*07-TNFα(-238)G-TNFα(-308)G with NTD, because the numbers were computed with statistical methods rather than obtained experimentally. Confirmation of this association requires the analysis of observed values, which is possible only in family studies.