The brain natriuretic peptide (BNP) precursor is the major immunoreactive form of BNP in patients with heart failure

Background: Peptides derived from brain natriuretic peptide (BNP) precursor (proBNP), BNP, and the N-terminal fragment of proBNP (NT-proBNP) are used as biomarkers of heart failure. It remains unclear which forms of these pepticles circulate in blood and which forms are measured by assays for these natriuretic peptides. Methods: To design assays for immunodetection of proBNP, NT-proBNP, and BNP, we used a panel of BNP- and NT-proBNP-specific monoclonal antibodies (MAbs). All MAbs were tested in 2-site combinations in time-resolved fluoroirnmunoassays with recombinant or synthetic antigens and plasma from heart failure (HF) patients. ProBNP and related molecules were assayed in HF plasma samples and plasma extracts by means of gel filtration fast protein liquid chromatography (FPLC) before and after protein fractionation on Sep-Pak C18 cartridges. Results: The limits of detection for BNP, proBNP, and NT-proBNP assays were 0.4,3, and 10 ng/L, respectively. Gel filtration-FPLC studies revealed 1 peak of NT-proBNP (similar to 25 kDa), 1 peak of proBNP (similar to 37 kDa), and 2 peaks of BNP immunoreactivity, a major peak (similar to 37 kDa) for proBNP, and a minor peak (similar to 4 kDa) for BNP. In patient plasma, the molar concentration of NT-proBNP was almost 10 times that of proBNP. The mean proBNP:BNP ratio in patient plasma was 6.3, ranging from 1.8 to 10.8. Conclusions: ProBNP is the major BNP-immunoreactive form in human blood. The proBNP:BNP ratio in plasma samples is dependent on the methods used for sample handling and for the measurement of the peptides. (c) 2007 American Association for Clinical Chemistry.