Radioimmunoassay for N-terminal probrain natriuretic peptide in human plasma

Abstract

Measurement of plasma levels of natriuretic peptides has been used to assess left ventricular dysfunction and prognosis. Recently levels of the N-terminal peptide fragment of the precursor of brain natriuretic peptide have been reported to be present in peripheral plasma and to be increased in chronic heart failure patients. Our aim in this study was to develop a radioimmunoassay for N-terminal proBNP, to compare its plasma concentrations in control subjects and in patients with end-stage heart failure and to define its relation to brain natriuretic peptide (BNP). A polyclonal antibody was raised in rabbits against human N-terminal proBNP fragment (amino acid 1-21). The plasma N-terminal proBNP concentrations were assayed directly without extraction. No detectable cross-reactivity existed with other natriuretic peptides: BNP, ANP or N-terminal proANP. The assay had a detection limit (2 SD from zero) of 9.7 pmol/L. Plasma N-terminal proBNP was 29 (13-75) (median (range)) pmol/L in the control group. There were no gender difference, male: 28 (13-61) vs. female 33 (13-75) pmol/L, p= NS, but there was a positive correlation to age (r=0.52, p<0.0001). In patients with end-stage heart failure the median N-terminal proBNP levels were increased significantly 616 (114-2781) pmol/L (p<0.001) and in pooled data N-terminal proBNP showed a close correlation to BNP (r=0.96, p<0.0001). Size exclusion of plasma extracts indicated that proBNP (1-108) may circulate both as intact prohormone and as split products, N-terminal proBNP (1-76) and BNP (77-108). Our results support the concept that N-terminal proBNP measurement could be a valuable tool in the biochemical indication of increased cardiac wall stress.

Biochemistry of Pro-B-Type Natriuretic Peptide-Derived Peptides: The Endocrine Heart Revisited

“…Immunoassays may nevertheless still detect the intact prohormone and therefore not reflect the actual bioactivity of the secreted peptides. In agreement with this suggestion, patients with congestive heart failure have increased plasma concentrations of immunoreactive N-terminal proBNP, and this immunoreactive material seems to also contain a polypeptide corresponding to the intact precursor (16,39,40 ). Thus, the congestion in heart failure patients may not be ameliorated by the secreted natriuretic peptides.…”

“…It is therefore important to realize that the preproBNP molecule does not exist in real life but is only as a theoretical concept. In contrast, proBNP is likely an existing small protein, the existence of which to date has been indicated by chromatographic studies and sequence-specific immunoassays (16,39,40 ). However, the actual precursor has not been identified on a molecular level, nor have the fragments thereof, apart from the C-terminal BNP-32 peptide itself (10 ).…”

“…These fragments are commonly referred to as "N-terminal proBNP", but the molecular heterogeneity also includes the intact precursor, in particular in heart failure patients (16,39,40,75 ). Cardiac secretion of proBNP and its N-terminal fragments has been demonstrated by blood sampling from the coronary sinus (76,77 ).…”

Quality Specifications for B-Type Natriuretic Peptide Assays

“…NT-proBNP assays theoretically should show 100% cross-reactivity with proBNP. Furthermore, intact proBNP has been described in human plasma, and oligomerization probably occurs (11 ). Assays for BNP and NT-proBNP may differ in their susceptibility to analytical interferences.…”

Clinical implications of normal B-type natriuretic peptide levels in patients with severe chronic heart failure

“…High BNP levels in patients with chronic heart failure (CHF) often indicate poor ventricular function and a worse prognosis, but studies have shown that in some patients with severe chronic heart failure the plasma BNP level is normal [8,9]. In clinical practice, with quality laboratory resources, BNP has been introduced to diagnosis, prognosis and as a guide for treatment [10][11][12][13][14][15]. In this study, 57 patients with severe CHF seen at our hospital from December 2002 to October 2009, were studied, focusing on plasma levels of BNP, clinical features and prognosis.…”