Objective To study the relationship between maternal thyroid function at each pregnancy trimester and neonatal screening results. Background Overt maternal thyroid dysfunction during gestation is associated with poor neonatal thyroid function. However, research on the relationship between suboptimal maternal thyroid function (assessed at three trimesters) and neonatal thyroid screening outcome is scarce. Design/Patients Prospective follow-up study during three trimesters of gestation in 886 Dutch Caucasian healthy pregnant women followed from 12-week gestation until term delivery (>37 weeks) and their neonates. Measurements The relation between neonatal data from the Congenital Hypothyroidism (CH) screening and maternal thyroid determinants [TSH, FT4 and thyroid peroxidase (TPO)-Ab] assessed at 12-, 24- and 36-week gestation. Results Boys have lower screening TT4 levels and their mothers have higher TSH levels at 24- and 36-week gestation. Higher maternal TSH levels (>97Æ5th percentile, as defined in 810 women without TPO-Ab at 12 weeks) at one or more times during pregnancy (O.R: 2Æ26, 95% CI: 1Æ20–4Æ29) and lower gestational age (O.R: 1Æ22, 95% CI: 1Æ05–1Æ41) are independently related to lower screening TT4 levels. Conclusions Maternal thyroid function during gestation is related to neonatal TT4 at screening. The finding of both lower neonatal TT4 levels in boys and higher TSH levels in mothers carrying boys is worthy of further investigation, as both observations may be meaningfully related.