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To determine a reference level for alphafetoprotein (AFP) in the amniotic fluid (AF) in pregnant women with insulin-dependent diabetes mellitus in order to suggest an explanation for the observed decrease in maternal serum AFP.Alpha-fetoprotein was measured in AF, maternal serum, or both in the second trimester in 287 pregnant women with insulin-dependent diabetes mellitus. Retrospectively, these AFP values were correlated with glycosylated hemoglobin (HbA1C) levels, early fetal growth delay, and congenital malformations.The median concentration of AFAFP was 0.89 multiples of the normative median (MoM) (n = 280; 95% confidence interval [CI] 0.88-0.93 MoM); the maternal serum AFP (MSAFP) value was 0.78 MoM (n = 155; 95% CI 0.77-0.84 MoM). A statistically significant but weak positive correlation was found between HbA1C and MSAFP (r2 = 0.033, P = .03), but the correlation with AFAFP was not statistically significant. The levels of AFP did not correlate with early fetal growth delay. One neural tube defect, but none of the 11 other major malformations, was detected by AFP screening.A physiologic basis for the decreased AFAFP and MSAFP levels is still obscure. Screening for congenital malformations in diabetic pregnancies should include both a mid-gestation ultrasound scan and MSAFP measurement. However, routine amniocentesis does not seem to be indicated. Concentrations of AFAFP may be corrected for the observed decrease.