Elevated total plasma homocysteine concentrations (tHcy), both fasting and post-methionine load, have been established as risk factors for vascular disease. Recently, we described the association of a 31 bp variable number of tandem repeats (VNTR) in the cystathionine beta-synthase (CBS) gene with both CBS enzyme activity and tHcy concentrations. In the present study, we determined the 31 bp VNTR genotypes in 2598 individuals of the Framingham Offspring Study and studied the association between this genotype and fasting, 2-h post-methionine load and delta (ie increase upon methionine loading) tHcy concentrations in 1416 subjects. We observed a positive association between the number of repeat units of the CBS 31 bp VNTR and both postload and delta tHcy concentrations. Adjustment for possible effect modifying factors like age, sex and vitamin (B6, B12 and folate) status did not change this observation. We hereby confirm the results of our earlier study, in which we found that this 31 bp VNTR is a genetic determinant of post-methionine load tHcy concentrations. Since also post-methionine load tHcy concentrations are found to be associated with an increased risk for cardiovascular disease (CVD), this 31 bp VNTR may be considered a risk factor for CVD.