Chemical and Biomolecular Engineering Department, University of California, Los Angeles, Los Angeles, CA 90095-1592, USA.
Biotechnol Appl Biochem. 2011 Jan-Feb;58(1):75-82. doi: 10.1002/bab.5.
A convenient homogeneous enzyme immunoassay for estradiol is described. Unlike heterogeneous immunoassays, which require time-consuming separation steps or expensive automated systems, homogeneous immunoassays, wherein all reagents are freely suspended in bulk solution, can be simple and fast without costly instrumentation. The key component of this assay system, an estradiol-reporter enzyme conjugate, was prepared by covalently binding β-estradiol-6-(O-carboxymethyl)oxime to glucose-6-phosphate dehydrogenase (G6PDH) by an N-hydroxysuccinimide-enhanced, carbodiimide-mediated coupling reaction. The estradiol-G6PDH activity can be repressed up to 46% upon anti-estradiol antibody binding. The lower detection limit of the assay is 1 nM estradiol in aqueous solution, and the standard curve is linear on logit-log scale-up to 6.7 µM estradiol. A detection limit of 11.5 nM in estradiol-spiked human serum samples suggests the feasibility of applying this assay to monitor estradiol levels for the prediction and prevention of ovarian hyperstimulation syndrome.
本文描述了一种简便的雌二醇均相酶免疫分析法。与需要耗时的分离步骤或昂贵的自动化系统的非均相免疫分析法不同，均相免疫分析法中所有试剂都可以在大量溶液中自由悬浮，无需昂贵的仪器即可简单快速。该测定系统的关键组分，即雌二醇报告酶结合物，是通过 N-羟基琥珀酰亚胺增强的、碳二亚胺介导的偶联反应将β-雌二醇-6-（O-羧甲基）肟共价结合到葡萄糖-6-磷酸脱氢酶（G6PDH）上制备的。抗雌二醇抗体结合可使雌二醇-G6PDH 活性抑制高达 46%。该测定法的检测下限为 1 nM 水溶液中的雌二醇，标准曲线在以对数-对数标度扩展至 6.7 µM 雌二醇时呈线性。在雌二醇加标人血清样品中的检测下限为 11.5 nM，表明该测定法可用于监测雌二醇水平，以预测和预防卵巢过度刺激综合征。
