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Hair concentrations are a noninvasive measure of cumulative antiretroviral exposure and the strongest predictor of viral suppression in large cohorts of nonpregnant patients. We examined hair concentrations of antiretrovirals in relation to virologic outcomes in pregnant and breastfeeding women for the first time.The Prevention of Malaria and HIV Disease in Tororo trial (NCT00993031) enrolled HIV-infected pregnant Ugandan women at 12–28 weeks gestation who were randomized to lopinavir or efavirenz-based antiretroviral therapy (ART). Small hair samples were collected at 30–34 weeks gestation and 10–25 weeks postpartum. Efavirenz and lopinavir hair concentrations were measured via liquid chromatography/tandem mass spectrometry. Multivariate logistic regression models examined predictors of viral suppression (HIV-1 RNA ≤400 copies/ml) at delivery and 24 weeks postpartum.Among 325 women, median CD4+ cell count was 366 cells/μl (interquartile range 270–488) at ART initiation. Mean self-reported 3-day adherence was greater than 97% in each arm. Viral suppression was achieved by 98.0% (efavirenz) and 87.4% (lopinavir) at delivery. At 24 weeks postpartum, 92.5% (efavirenz) and 90.6% (lopinavir) achieved viral suppression; 88% of women were breastfeeding. In multivariate models including self-reported adherence and pretreatment HIV-1 RNA, antiretroviral hair concentrations were the strongest predictor of viral suppression at delivery [efavirenz: adjusted odds ratio (aOR) 1.86 per doubling in concentration, 95% confidence interval (CI) 1.14–3.1, P = 0.013; lopinavir: aOR 1.90, 95% CI 1.33–2.7, P = 0.0004] and 24 weeks postpartum (efavirenz: aOR 1.81, 95% CI 1.22–2.7, P = 0.003; lopinavir: aOR 1.53, 95% CI 1.05–2.2, P = 0.026).Antiretroviral hair concentrations represent an innovative tool that strongly predicts viral suppression among HIV-infected childbearing women during the critical periods of delivery and breastfeeding.