Objective: To evaluate the ability of maternal serum-free β-human chorionic gonadotrophin (β-hCG), pregnancy-associated plasma protein-A (PAPP-A), and alpha fetoprotein (AFP) levels in the screening of fetuses with hemoglobin (Hb) Bart's disease among pregnancies at risk. Materials and Methods: Pregnancies at risk for fetal Hb Bart's disease scheduled for cordocentesis at 18 to 22 weeks were recruited into the study. Maternal serum-free β-hCG, PAPP-A, and AFP concentrations were measured before cordocentesis, and the final fetal diagnosis of Hb Bart disease was based on fetal Hb typing using high-performance liquid chromatography. Results: Of 57 recruited pregnancies, 11 had fetal Hb Bart's disease and 46 were unaffected. Maternal serum alpha-fetoprotein (MSAFP) concentrations were significantly higher in women with fetal Hb Bart's disease than those with unaffected fetuses (median 99.53 vs 50.83, P < 0.001), whereas the concentrations of free β-hCG and PAPP-A were not significantly different between the two groups (P = 0.543 and 0.777, respectively). Conclusion: Second-trimester MSAFP may be clinically a useful screening test for fetal Hb Bart's disease among pregnancies at risk. © 2011 John Wiley & Sons, Ltd.