Education Ministry Key Laboratory on Luminescence and Real-Time Analysis, College of Chemistry and Chemical Engineering, Southwest University, Chongqing 400715, People's Republic of China.
Biosens Bioelectron. 2013 Dec 15;50:325-30. doi: 10.1016/j.bios.2013.06.050. Epub 2013 Jul 4.
In this work, a new electrochemical aptasensor based on direct electron transfer and electrocatalysis of horseradish peroxidase (HRP) using exonuclease-catalyzed target recycling and hybridization chain reaction (HCR) for signal amplification was developed for highly sensitive detection of thrombin. The electrochemical signal was originated from HRP without the addition or labeling of redox probes. To construct the aptasensor, the capture probe was immobilized on gold nanoparticles (AuNPs) modified electrode for the following hybridization with the complementary thrombin binding aptamer. In the presence of thrombin, the formation of aptamer-thrombin complex would result in the dissociation of aptamer from the double-strand DNA (dsDNA). Subsequently, with the employment of exonuclease, aptamer was selectively digested and thrombin could be released for analyte recycling. The capture probe and two hairpin helper DNAs lead to the formation of extended dsDNA polymers through HCR on the electrode surface. Then the biotin-labeled dsDNA polymers could introduce numerous avidin-labeled HRP, resulting in significantly amplified electrochemical signal through the direct electrochemistry and electrocatalysis of HRP. The proposed strategy combined the amplification of analyte recycling and HCR, as well as the inherent electroactivity and catalytic activity of HRP, which exhibited high sensitivity for thrombin determination with an ultra-low detection limit of 1.2×10(-13) M. Moreover, the detection scheme could be easily extended to the detection of other biomolecules.
在这项工作中，开发了一种基于辣根过氧化物酶（HRP）的直接电子转移和电催化的新型电化学适体传感器，该传感器利用外切酶催化的靶标循环和杂交链式反应（HCR）进行信号放大，用于高灵敏检测凝血酶。电化学信号源自 HRP，无需添加或标记氧化还原探针。为了构建适体传感器，捕获探针被固定在金纳米粒子（AuNPs）修饰的电极上，用于与互补的凝血酶结合适体进行杂交。在凝血酶存在的情况下，适体-凝血酶复合物的形成会导致适体从双链 DNA（dsDNA）中解离。随后，利用外切酶，适体被选择性地消化，凝血酶可以被释放进行分析物的循环。捕获探针和两条发夹辅助 DNA 通过在电极表面上的 HCR 形成扩展的 dsDNA 聚合物。然后，生物素标记的 dsDNA 聚合物可以引入大量的亲和素标记的 HRP，通过 HRP 的直接电化学和电催化产生显著放大的电化学信号。所提出的策略结合了分析物循环和 HCR 的放大，以及 HRP 的固有电化学活性和催化活性，用于凝血酶检测具有超低检测限 1.2×10(-13) M。此外，该检测方案可以很容易地扩展到其他生物分子的检测。
