Background: Folate is involved in DNA synthesis and methylation and may thereby influence carcinogenesis. Objectives: We examined plasma folate (P-folate) concentration in relation to genotypes of the folate-metabolizing enzyme methylenetetrahydrofolate reductase [MTHFR 677C [right arrow] T (rs 1801133) and 1298A [right arrow] C (rs1801131)]. We also explored whether P-folate was associated with risk of postmenopausal breast cancer overall and in subgroups with genetic variants of the MTHFR single nucleotide polymorphisms (SNPs). Design: This nested case-control study included 313 cases (age 55-73 y at baseline) with invasive breast cancer and 626 control subjects, matched on age and blood-sample date, from the population-based Malmo Diet and Cancer cohort. P-folate and MTHFR genotypes were determined for 310 cases and 611 controls. P-folate according to genotype was calculated by using analysis of variance. Odds ratios were obtained by using logistic regression. All tests were 2-sided. Results: The variant 677T allele was associated with lower P-folate. In women with the 677T allele, a high P-folate concentration was associated with increased breast cancer risk (P for trend across P-folate tertiles = 0.03). Interaction was seen between the 677C [right arrow] T SNP and P-folate (P = 0.002). A positive association, which was seen between P-folate and breast cancer risk in 1298AA women (P = 0.01), was probably due to linkage between the 2 SNPs. Overall, and in women with other genotypes, no significant associations were observed. Conclusions: Our results suggest an association of high P-folate concentration with increased risk of postmenopausal breast cancer in carriers of the 677T allele. The findings underline the importance of genetic variation of MTHFR in the complex relation between folate and cancer. doi: 10.3945/ajcn.2009.28064.