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Background: The National Children's Study (NCS) Vanguard Study began enrollment in January 2009 as an initial pilot study for a planned large-scale, longitudinal U.S. cohort study of the effect of environmental influences on child health and development, with biological and environmental sample collection conducted in seven locations from April 2009 to October 2010. We sought to determine rates of consent for, and success of collection of, maternal and paternal biospecimens before and during pregnancy in the NCS Vanguard Study., Methods: Samples of blood, saliva, vaginal swabs, urine, hair, and nails were collected before and during pregnancy. All specimens were sent to a central repository for processing, storage, and quality assessment., Results: Of 780 pregnant women asked to consent to sample collection, 773 (>99%) agreed, and of 295 nonpregnant women, 292 (99%) agreed. Of 440 fathers asked to consent to sample collection, 435 (99%) agreed. Frequency of successful collection of biospecimens varied depending on sample and visit type. In descending order, the ranges over all visit types of the proportion of expected samples successfully collected from women were: urine, 92.5 to 95.7%; hair, 89.6 to 92.5%; vaginal swab, 84.2 to 88.5%; blood, 74.9 to 78.5%; 2-day saliva, 65.8 to 81.6%; and nails, 76.4 to 76.7%. For fathers, rates were highest for urine (94.9%) and lowest for hair (63.0%)., Conclusion: High rates of consent for and collection of a wide variety of biospecimens can be achieved in prospective epidemiologic cohort studies of pregnant women. Ease of sample collection may be a primary factor influencing successful biospecimen collection., (© 2015 Wiley Periodicals, Inc.)