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首页> 外文期刊>Analytical chemistry >Magnetic Electrochemical Sensing Platform for Biomonitoring of Exposure to Organophosphorus Pesticides and Nerve Agents Based on Simultaneous Measurement of Total Enzyme Amount and Enzyme Activity
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Magnetic Electrochemical Sensing Platform for Biomonitoring of Exposure to Organophosphorus Pesticides and Nerve Agents Based on Simultaneous Measurement of Total Enzyme Amount and Enzyme Activity

机译：基于同时测量总酶量和酶活性的生物电化学监测有机磷农药和神经制剂的电磁电化学传感平台

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摘要

We report a new approach for electrochemical quantification of enzymatic inhibition and phosphorylation for biomonitoring of exposure to organophosphorus (OP) pesticides and nerve agents based on a magnetic bead (MB) immunosensing platform. The principle of this approach is based on the combination of MB immunocapture-based enzyme activity assay and competitive immunoassay of the total amount of enzyme for simultaneous detection of enzyme inhibition and phosphorylation in biological fluids. Butyrylcholinesterase (BChE) was chosen as a model enzyme. In competitive immunoassay, the target BChE in a sample competes with the BChE immobilized on the MBs to bind to the limited sites of anti-BChE antibody labeled with quantum dots (QD-anti-BChE), followed by stripping voltammetric analysis of the bound QD conjugate on the MBs. This assay shows a linear response over the total BChE concentration range of 0.1-20 nM. Simultaneous real time BChE activity was measured on an electrochemical carbon nanotube-based sensor coupled with a microflow injection system after immunocapture by the MB-anti-BChE conjugate. Therefore, the formed phosphorylated BChE adduct (OP-BChE) can be estimated by the difference values of the total amount of BChE (including active and OP-inhibited) and active BChE from established calibration curves. This approach not only eliminates the difficulty in screening of low-dose OP exposure (less than 20percent inhibition of BChE) because of individual variation of BChE values but also avoids the drawback of the scarce availability of OP-BChE antibody. It is sensitive enough to detect 0.5 nM OP-BChE, which is less than 2percent BChE inhibition. This method offers a new method for rapid, accurate, selective, and inexpensive quantification of OP-BChE and enzyme inhibition for biomonitoring of OP and nerve agent exposures.
机译：我们报告了一种新的电化学抑制和磷酸化的化学定量方法，用于基于磁珠（MB）免疫传感平台的有机磷（OP）农药和神经毒剂生物暴露监测。这种方法的原理是基于MB免疫捕获的酶活性测定法和总酶竞争性免疫测定法的结合，用于同时检测生物液中的酶抑制和磷酸化。选择丁酰胆碱酯酶（BChE）作为模型酶。在竞争性免疫测定中，样品中的目标BChE与固定在MBs上的BChE竞争，以结合用量子点标记的抗BChE抗体的有限位点（QD-anti-BChE），然后对结合的QD进行伏安分析在MB上共轭。该测定法显示了在0.1-20 nM的总BChE浓度范围内的线性响应。在通过MB-抗BChE偶联物免疫捕获后，在结合了微流注射系统的电化学碳纳米管基传感器上测量了实时实时BChE活性。因此，可以根据已建立的校准曲线通过BChE总量（包括活性和OP抑制）与活性BChE的差值估算形成的磷酸化BChE加合物（OP-BChE）。这种方法不仅消除了由于BChE值的个体差异而导致的低剂量OP暴露筛查（小于20％的BChE抑制）的困难，而且避免了OP-BChE抗体稀缺的缺点。它足够灵敏，可以检测到0.5 nM OP-BChE，其抑制率不到2％。该方法为快速，准确，选择性和廉价地定量OP-BChE和酶抑制提供了一种新方法，可用于生物监测OP和神经制剂的暴露。

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