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This prospective study investigates whether amino terminal proB-type natriuretic peptide (NT-proBNP) levels are elevated in neonates with congenital heart defect (CHD). NT-proBNP levels in the umbilical cord blood of 60 neonates with prenatally diagnosed CHD, in the cord blood of 200 control subjects, and in the plasma of the respective mothers were analyzed using an automated enzyme immunoassay. NT-proBNP concentrations in the cord blood of the CHD group were significantly elevated compared with controls median (range): 158 pg/mL (378–3631 pg/mL) and 626 pg/mL (153–2518 pg/mL); pvalue <0.001. The NT-proBNP concentrations of the newborns and their mothers did correlate neither in the CHD nor in the control group. In 54 patients with CHD, NT-proBNP levels were measured on the median 5th day of life. They showed a significant increase (median 1665 pg/mL and 19222 pg/mL; p< 0.001). NT-proBNP levels in the cord blood of neonates with CHD are significantly elevated and show a marked increase in the first week of life. Furthermore, this study confirms previously published reference intervals of NT-proBNP in the cord blood of healthy neonates as well as the finding that there is no exchange of NT-proBNP across the placenta.