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Dual Enzymatic Detection by Bulk Electrogenerated Chemiluminescence

机译:本体电化学发光双重酶检测

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The combination of enzymes, as recognition elements for specific analytes, and of electrogenerated chemiluminescence (ECL) as a readout method has proven to be a valuable strategy for sensitive and specific analytical detection. However, ECL is intrinsically a 2D process which could potentially limit the analysis of inhomogeneous samples. Here, we show how a bulk ECL signal, generated by thousands of carbon microbeads remotely addressed via bipolar electrochemistry, are implemented as a powerful tool for the concomitant ECL sensing and imaging of two enzymatic substrates. We selected two enzymes (glucose dehydrogenase and choline oxidase) that react with their respective model substrates and produce in situ chemical species (beta-nicotinamide adenine dinucleotide (NADH) and H2O2) acting as coreactants for the ECL emission of different luminophores ([Ru(bpy)(3)](2+) at lambda = 620 nm and luminol at lambda = 425 nm, respectively). Both enzymes are spatially separated in the same capillary. We demonstrate thus the simultaneous quantitative determination of both glucose and choline over a wide concentration range. The originality of this remote approach is to provide a global chemical view through one single ECL image of inhomogeneous samples such as a biochemical concentration gradient in a capillary configuration. Finally, we report the first proof-of-concept of dual biosensing based on this bulk ECL method for the simultaneous imaging of both enzymatic analytes at distinct wavelengths.
机译:酶(作为特定分析物的识别元素)与电化学发光(ECL)作为一种读出方法的结合已被证明是用于灵敏和特异性分析检测的有价值的策略。但是,ECL本质上是2D过程,可能会限制非均质样品的分析。在这里,我们展示了如何通过双极性电化学方法远程解决由成千上万个碳微珠产生的大量ECL信号,将其作为同时进行ECL感测和两种酶促底物成像的强大工具。我们选择了两种酶(葡萄糖脱氢酶和胆碱氧化酶),它们分别与各自的模型底物反应,并产生原位化学物种(β-烟酰胺腺嘌呤二核苷酸(NADH)和H2O2)充当不同发光体的ECL发射的共反应物([Ru( bpy)(3)](2+)分别在λ= 620 nm和鲁米诺在λ= 425 nm)。两种酶在同一毛细管中在空间上分开。因此,我们证明了在宽浓度范围内同时定量测定葡萄糖和胆碱的能力。这种远程方法的独创性是通过不均匀样品的单个ECL图像(例如毛细管结构中的生化浓度梯度)来提供全局化学视图。最后,我们报告了基于这种批量ECL方法的双重生物传感的第一个概念验证,用于同时在不同波长下对两种酶分析物进行成像。

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