A Novel Immunological Assay for Hepcidin Quantification in Human Serum
A Novel Immunological Assay for Hepcidin Quantification in Human Serum
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DOI：
10.1371/journal.pone.0004581
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发表时间：
2009-02-24
期刊：
影响因子：
3.7
通讯作者：
Mamalaki, Avgi
中科院分区：
文献类型：
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作者：
Koliaraki, Vasiliki;Marinou, Martha;Mamalaki, Avgi
背景：Hepcidin是一种富含25个氨基酸半胱氨酸的铁调节肽。hepcidin浓度升高导致巨噬细胞中的铁隔离，促进慢性疾病贫血的发病机制，而hepcidin浓度降低则在铁缺乏和原发性铁超载疾病（如遗传性血色素沉着症）中观察到。人类血液或尿液中的Hepcidin定量可能为铁稳态紊乱的发病机制提供进一步的见解，并可能为临床医生鉴别诊断贫血提供有价值的工具。本研究描述了一种特异性且不需要操作人员的免疫测定法，用于人血清中hepcidin的定量。方法和发现：采用重组hepcidin25-His肽段和针对该肽段的多克隆抗体，建立了ELISA法测定hepcidin血清浓度，该抗体能够识别天然hepcidin。ELISA法检测范围为10 ~ 1500 μ g/L，检出限为5.4 μ g/L。试验内和试验间方差系数分别为8-15%和5-16%。平均线性和回收率分别为101%和107%。7例少年血色素沉着症患者（12.8 μ g/L）和10例缺铁性贫血患者（15.7 μ g/L）的平均hepcidin水平显著低于32例年龄匹配的健康对照（42.7 μ g/L）， 7例霍奇金淋巴瘤患者（116.7 μ g/L）的平均hepcidin水平显著高于32例年龄匹配的健康对照（42.7 μ g/L）。结论：我们描述了一种新的测定人血清中hepcidin的简单ELISA方法，具有足够的准确性和重复性。
Background: Hepcidin is a 25-aminoacid cysteine-rich iron regulating peptide. Increased hepcidin concentrations lead to iron sequestration in macrophages, contributing to the pathogenesis of anaemia of chronic disease whereas decreased hepcidin is observed in iron deficiency and primary iron overload diseases such as hereditary hemochromatosis. Hepcidin quantification in human blood or urine may provide further insights for the pathogenesis of disorders of iron homeostasis and might prove a valuable tool for clinicians for the differential diagnosis of anaemia. This study describes a specific and non-operator demanding immunoassay for hepcidin quantification in human sera.Methods and Findings: An ELISA assay was developed for measuring hepcidin serum concentration using a recombinant hepcidin25-His peptide and a polyclonal antibody against this peptide, which was able to identify native hepcidin. The ELISA assay had a detection range of 10-1500 mu g/L and a detection limit of 5.4 mu g/L. The intra-and interassay coefficients of variance ranged from 8-15% and 5-16%, respectively. Mean linearity and recovery were 101% and 107%, respectively. Mean hepcidin levels were significantly lower in 7 patients with juvenile hemochromatosis (12.8 mu g/L) and 10 patients with iron deficiency anemia (15.7 mu g/L) and higher in 7 patients with Hodgkin lymphoma (116.7 mu g/L) compared to 32 age-matched healthy controls (42.7 mu g/L).Conclusions: We describe a new simple ELISA assay for measuring hepcidin in human serum with sufficient accuracy and reproducibility.