In this study the impact of pregnancy duration on the measured level of HSV-2 antibodies was assessed. The study population comprised 35940 pregnant women in Norway, in 1992–4, followed during pregnancy. A random sample of 960 women was selected. A mean of 2·6 serum samples from each woman were analysed for HSV-2 specific IgG antibodies at different times in pregnancy. Crude and adjusted odds ratios were estimated in logistic regression models taking all observations per women into account. Twenty-seven percent of the pregnant women had antibodies against HSV-2 in the first trimester. The adjusted odds ratio of being HSV-2 antibody positive decreased during the pregnancy and was 0·5 (0·2–0·9, 95% confidence interval) in the 40th as compared to the 10th week of pregnancy. About 50% of initially HSV-2 positive women did not have detecable antibodies by the end of the pregnancy. This may be explained by haemodilution during pregnancy. Our findings have diagnostic implications and should encourage further studies.