Seferian Karina R, Tamm Natalia N, Semenov Alexander G, Mukharyamova Kadriya S, Tolstaya Anastasya A, Koshkina Ekaterina V, Kara Andrei N, Krasnoselsky Mihail I, Apple Fred S, Esakova Tatiana V, Filatov Vladimir L, Katrukha Alexey G
HyTest Ltd., Turku, Finland.
Clin Chem. 2007 May;53(5):866-73. doi: 10.1373/clinchem.2006.076141. Epub 2007 Mar 23.
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.
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.
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.
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.
源自脑钠肽（BNP）前体（proBNP）、BNP以及proBNP的N端片段（NT-proBNP）的肽类被用作心力衰竭的生物标志物。目前尚不清楚这些肽类的哪些形式在血液中循环，以及哪些形式可通过这些利钠肽检测法进行检测。
为设计用于免疫检测proBNP、NT-proBNP和BNP的检测法，我们使用了一组BNP和NT-proBNP特异性单克隆抗体（MAb）。所有MAb均在时间分辨荧光免疫分析中与重组或合成抗原以及心力衰竭（HF）患者的血浆进行双位点组合测试。在Sep-Pak C18柱上进行蛋白质分级分离前后，通过凝胶过滤快速蛋白质液相色谱（FPLC）对HF血浆样本和血浆提取物中的proBNP及相关分子进行检测。
BNP、proBNP和NT-proBNP检测法的检测限分别为0.4、3和10 ng/L。凝胶过滤-FPLC研究显示NT-proBNP有1个峰（约25 kDa），proBNP有1个峰（约37 kDa），BNP免疫反应性有2个峰，一个主要峰（约37 kDa）对应proBNP，一个次要峰（约4 kDa）对应BNP。在患者血浆中，NT-proBNP的摩尔浓度几乎是proBNP的10倍。患者血浆中proBNP与BNP的平均比值为6.3，范围在1.8至10.8之间。
proBNP是人类血液中主要的BNP免疫反应形式。血浆样本中proBNP与BNP的比值取决于样本处理方法和肽类的测量方法。
