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Objectives: To compare plasma BNP values determined by a conventional-type and a novel-type of BNP assays., Design and Methods: Plasma samples (n=94) from HF patients were analyzed by the novel-type "Single Epitope Sandwich" (SES assay prototype) and the conventional-type Siemens ADVIA Centaur BNP assays. Both assays were calibrated using recombinant proBNP (expressed in E. coli)., Results: The SES assay measured 1.2- to 7.2-fold (2.1±0.9; mean, SD) greater BNP concentrations compared to the Siemens assay. A subset of six samples (6.4%) demonstrated the largest (3- to 7.2-fold higher) difference., Conclusions: The SES prototype assay appears to more accurately measure the absolute concentrations of BNP immunoreactive forms., (Copyright © 2010 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.)

Background: Brain natriuretic peptide (BNP) is an unstable molecule that can rapidly lose immunologic activity in blood. Conventional sandwich BNP immunoassays use 2 antibodies specific to 2 different epitopes. Larger distances between epitopes are associated with a greater probability of proteolysis sites being located between the antibody-binding sites, and thus such assays have an increased susceptibility to underdetect BNP because of the increased likelihood of proteolytic degradation. The purpose of our study was to develop a sandwich immunoassay for the precise quantification of BNP and BNP precursor (proBNP) in human blood that is not susceptible to proteolysis., Methods: Mice were immunized with an immune complex consisting of monoclonal antibody (MAb) 24C5 (specific for BNP peptide 11-22) and the entire BNP molecule. The MAb used in our assay (Ab-BNP2) recognizes the immune complex but neither free BNP nor MAb 24C5., Results: We used MAbs 24C5 and Ab-BNP2 to develop a new type of sandwich BNP assay (the "single-epitope sandwich assay"), which requires only a short BNP fragment (fragment 11-22) for immunodetection. This assay recognizes both BNP and proBNP with the same efficiency and sensitivity and demonstrates both considerably less susceptibility to antigen degradation and greater stability of the measured antigen than conventional sandwich BNP immunoassays., Conclusions: We have developed this sensitive single-epitope sandwich assay for detecting BNP, proBNP, and their fragments in human blood. This assay appears promising for use in clinical studies to assist in triage, management, and outcomes assessment in heart failure patients.

Background: Brain natriuretic peptide (BNP) or NT-proBNP (N-terminal fragment of BNP precursor) measurements are recommended as aids in diagnosis and prognosis of patients with heart failure. Recently it has been shown that proBNP is O-glycosylated in human blood. The goal of this study was to map sites on the NT-proBNP molecule that should be recognized by antibodies used in optimal NT-proBNP assays., Methods: We analyzed endogenous NT-proBNP by several immunochemical methods using a broad panel of monoclonal antibodies specific to different epitopes of the NT-proBNP molecule., Results: Treatment of endogenous NT-proBNP by a mixture of glycosidases resulted in significant improvement of the interaction between deglycosylated NT-proBNP and monoclonal antibodies (MAbs) specific to the mid-fragment of the molecule. MAbs specific to the N- and C-terminal parts of NT-proBNP (epitopes 13-24 and 63-76) were able to recognize glycosylated and deglycosylated protein with similar efficiency., Conclusions: The central part of endogenous NT-proBNP is glycosylated, making it almost "invisible" for the antibodies specific to the mid-fragment of the molecule. Thus sandwich assays using even one antibody (poly- or monoclonal) specific to the central part of the molecule could underestimate the real concentration of endogenous NT-proBNP. MAbs specific to the N- and C-terminal parts of NT-proBNP (epitopes 13-24 and 63-76) are the best candidates to be used in an assay for optimal NT-proBNP immunodetection.

Introduction: Brain natriuretic peptide (BNP) is produced by the ventricles of the heart and is a biomarker for heart failure. Several commercial assays are now available. We evaluated the performance characteristics of the ARCHITECT BNP assay., Methods: We evaluated the limit of blank, limit of detection, linearity and imprecision. Method comparison studies were performed with 3 other automated BNP assays including the ADVIA Centaur, AxSYM, and UniCel DxI 800 methods., Results: The mean LOB and LOD of the Architect assay were 3.5 and 5.8 ng/L, respectively. Imprecision studies yielded within run CVs of 1.1 to 5.1% and total CVs of 2.3 to 5.3% using human plasma based multi-constituent controls at concentrations of 92, 500, and 3500 ng/L. The maximum deviation from the target recovery for dilution linearity was 9.6%. Concordance with other BNP assays at a 100 ng/l cutoff was 91 to 98% and kappa statistics were 0.78 to 0.96. The mean difference between the Architect and Advia Centaur methods was positive. For the other methods, the mean difference with the Architect was negative., Conclusions: The Architect BNP assay shows good performance characteristics with total imprecision < or =5.3%. It agrees well with the Advia Centaur, AxSYM, and UniCel DxI BNP assays.

Background: Measurement of amino-terminal pro-B-type natriuretic peptide (NT-proBNP) is useful for evaluating patients with heart failure (HF)., Methods: We evaluated the performance of a new automated NT-proBNP assay., Results: The VITROS NT-proBNP assay had mean within-run and total imprecision of 1.0% and 3.4% at NT-proBNP concentrations from 67-27,500 ng/l. Acceptable linearity, functional/analytical sensitivity were demonstrated. Anticoagulant/tube types had no effect on results. Excellent sample stability and no high-dose hook were observed. High correlation between the VITROS and Elecsys methods was demonstrated (r=0.995; P<.001), with 98.3% clinical concordance. VITROS NT-proBNP concentrations were significantly higher in HF subjects than those without (1210 versus 68 ng/l; P<.001) and associated with HF symptom severity (P<.001). The VITROS assay had AUC for HF of 0.95 (P<.001), and had excellent NPV for excluding HF., Conclusions: The automated VITROS NT-proBNP assay demonstrates excellent analytical and clinical performance for evaluating the presence and severity of HF.

Background: NT-proBNP measurements aid in the evaluation of patients with suspected heart failure (HF) and may facilitate risk stratification in patients with HF and acute coronary syndrome (ACS). Point-of-care (POC) assays may provide more timely results and potentially improve patient outcomes., Methods: We evaluated the analytical performance of the Response Biomedical Corporation whole blood RAMP amino-terminal pro-B type natriuretic peptide (NT-proBNP) POC assay compared to the Roche Elecsys proBNP (NT-proBNP) assay., Results: Intra-day and total imprecision (% CV) ranged from 5.5% to 10.3% at 140, 449 and 1675 ng/L. The lowest concentration that yields a 20% CV was 57 ng/L. The lower limit of detection was 18 ng/L. The upper limit of linearity was validated to 23,428 ng/L with an average recovery of 95%. Correlation by Passing and Bablok regression yielded RAMP=1.01 Elecsys+14.6, r=0.98 (n=540; range of Elecsys values <5 to >35,000). Concordance of RAMP versus Elecsys using cut-offs of 125 ng/L for subjects <75 years and 450 ng/L for subjects > or =75 was 92% (95% CI 89-94%) for a group consisting of 127 apparently healthy individuals and 208 non-healthy subjects without HF, and 99% (95% CI 97-100%) for patients with HF, using the New York Heart Association (NYHA) functional classification. Overall, 80%, 87%, 97% and 100% of the RAMP results and 77%, 85%, 96% and 100% of the Elecsys results were greater than the age appropriate cut-off for NYHA I, II, III or IV groups. For both the RAMP and Elecsys results, the median NT-proBNP value was statistically correlated (increasing) with NYHA I, II, III or IV groups, respectively (p<0.0001), with no significant difference between the two methods., Conclusions: The POC Response Biomedical RAMP NT-proBNP assay provides comparable results that measured on the FDA cleared Roche Elecsys central laboratory platform.

Background: Further investigations are warranted to better characterize variables that may confound the clinical interpretation of plasma natriuretic peptide measurements, which are increasingly recognized to have diagnostic and predictive importance., Methods: Blood samples (EDTA plasma) from patients (n = 206) attending clinics for the medical treatment and follow-up of obesity were analyzed for B-type natriuretic peptide (BNP; Bayer assay) and the N-terminal segment of its prohormone (NT-proBNP; Roche assay). Natriuretic peptide concentration ranges were evaluated in those without diagnosis of congestive heart failure (CHF) or chronic kidney disease (CKD)., Results: BNP and NT-proBNP were directly correlated (r = 0.87; P = 0.01), with NT-proBNP concentrations higher relative to BNP. Of obese patients without CHF or CKD, 21.6% (40 of 185) had NT-proBNP concentrations greater than the published assay upper reference limit. Concentrations of both natriuretic peptides were higher in patients currently exposed to beta blockers, patients with the diagnosis of hypertension or type 2 diabetes, and patients with a history of gastric bypass surgery. An inverse relationship between body mass index (BMI) and both BNP and NT-proBNP was evident. According to the National Institutes of Health, National Heart, Lung, and Blood Institute classification, more than 95% of the participants sampled in this study were either obesity class 2 (35 kg/m(2) < BMI < 39.9 kg/m(2)) or class 3 (BMI >or=40 kg/m(2)), Conclusions: A substantial proportion of obese patients without CHF or CKD have concentrations greater than the upper reference limit for NT-proBNP but not for simultaneously measured BNP. A history of gastric bypass surgery appeared to be a significant predictor of increased natriuretic peptide concentrations when assessed in a population of patients with class 2 or 3 obesity.

Background: In patients with end-stage renal disease (ESRD), the ability of single and multiple biomarker monitoring to predict adverse outcomes has not been well established. This study determined the prognostic value of multiple biomarkers for all-cause death over 2 years in 399 ESRD patients., Methods: The risk of all-cause death was determined by use of multiple biomarkers based on concentrations for a reference population (normal) and cutoffs based on tertile distributions in the ESRD group. Biomarkers studied included N-terminal pro-B-type natriuretic peptide (NT-proBNP), high-sensitivity C-reactive protein (hsCRP; Dade Behring and Roche assays), and cardiac troponin T (cTnT; Roche) and I (cTnI; Dade Behring and Beckman Coulter assays). Relative risks of death were estimated and survival curves computed., Results: A total of 101 deaths occurred during 594 patient-years of follow-up. Increased NT-proBNP concentrations were not predictive of death on the basis of the normal cutoffs. However, tertile analysis of NT-proBNP was significantly predictive of death and had a ROC area under the curve equivalent to or better than any of the other biomarkers. Biomarkers independently predictive of survival were hsCRP (P <0.001, either assay), cTnT (P <0.05), and cTnI (Dade, P <0.05). Two-year mortality rates were 6% (n = 45) with normal hsCRP, cTnI, and cTnT concentrations; 19% (n = 173) with increased hsCRP or cTnT and normal cTnI; 44% (n = 160) with both hsCRP and cTnT increased and normal cTnI; 61% (n = 21) with increased cTnI (Dade) or 47% (n = 74) with increased cTnI (Beckman) regardless of hsCRP or cTnT concentrations. Defined by the normal cutoffs, increased concentrations of biomarkers were present in various proportions of the 399 patients with ESRD: NT-proBNP, 99%; hsCRP, 46% (both Roche and Dade assays); cTnT, 85%; cTnI, 19% (Beckman assay) and 5% (Dade assay)., Conclusions: Although mechanisms likely vary for causation, increased plasma hsCRP, cTnT, and cTnI above the cutoffs for our reference (normal) population were all independently predictive of subsequent death in ESRD patients. Tertile analysis for NT-proBNP also demonstrated prognostic value.