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 peptides 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 fluoroimmunoassays 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 (approximately 25 kDa), 1 peak of proBNP (approximately 37 kDa), and 2 peaks of BNP immunoreactivity, a major peak (approximately 37 kDa) for proBNP, and a minor peak (approximately 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.
Seferian
Karina R
KR
HyTest Ltd., Turku, Finland.
Tamm
Natalia N
NN
Semenov
Alexander G
AG
Mukharyamova
Kadriya S
KS
Tolstaya
Anastasya A
AA
Koshkina
Ekaterina V
EV
Kara
Andrei N
AN
Krasnoselsky
Mihail I
MI
Apple
Fred S
FS
Esakova
Tatiana V
TV
Filatov
Vladimir L
VL
Katrukha
Alexey G
AG
D000911:Antibodies, Monoclonal; D015415:Biomarkers / Q000097:blood; D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D015200:Fluoroimmunoassay; D006333:Heart Failure / Q000097:blood*; D006801:Humans; D020097:Natriuretic Peptide, Brain / Q000097:blood* / Q000276:immunology; D010446:Peptide Fragments / Q000097:blood* / Q000276:immunology; D011498:Protein Precursors / Q000097:blood* / Q000276:immunology; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D015415:Biomarkers; D010446:Peptide Fragments; D011498:Protein Precursors; C109794:pro-brain natriuretic peptide (1-76); D020097:Natriuretic Peptide, Brain