Key Laboratory of Eco-environments in the Three Gorges Reservoir Region (Ministry of Education), School of Chemistry and Chemical Engineering, Southwest University, Chongqing 400715, China.
Anal Biochem. 2013 Oct 15;441(2):95-100. doi: 10.1016/j.ab.2013.07.023. Epub 2013 Jul 26.
A sensitive chronocoulometric aptasensor for the detection of thrombin has been developed based on gold nanoparticle amplification. The functional gold nanoparticles, loaded with link DNA (LDNA) and report DNA (RDNA), were immobilized on an electrode by thrombin aptamers performing as a recognition element and capture probe. LDNA was complementary to the thrombin aptamers and RDNA was noncomplementary, but could combine with [Ru(NH₃)₆]³⁺ (RuHex) cations. Electrochemical signals obtained by RuHex that bound quantitatively to the negatively charged phosphate backbone of DNA via electrostatic interactions were measured by chronocoulometry. In the presence of thrombin, the combination of thrombin and thrombin aptamers and the release of the functional gold nanoparticles could induce a significant decrease in chronocoulometric signal. The incorporation of gold nanoparticles in the chronocoulometric aptasensor significantly enhanced the sensitivity. The performance of the aptasensor was further increased by the optimization of the surface density of aptamers. Under optimum conditions, the chronocoulometric aptasensor exhibited a wide linear response range of 0.1-18.5 nM with a detection limit of 30 pM. The results demonstrated that this nanoparticle-based amplification strategy offers a simple and effective approach to detect thrombin.
基于金纳米粒子放大作用，研制了一种用于检测凝血酶的灵敏计时库仑型适体传感器。功能化的金纳米粒子，负载链接 DNA（LDNA）和报告 DNA（RDNA），通过作为识别元件和捕获探针的凝血酶适体固定在电极上。LDNA 与凝血酶适体互补，而 RDNA 则不互补，但可以与[Ru（NH₃）₆]³⁺（RuHex）阳离子结合。通过计时库仑法测量通过静电相互作用定量结合到 DNA 带负电荷的磷酸骨架上的 RuHex 的电化学信号。在凝血酶存在下，凝血酶与凝血酶适体的结合以及功能化金纳米粒子的释放会导致计时库仑信号显著降低。在计时库仑适体传感器中加入金纳米粒子显著提高了灵敏度。通过优化适体的表面密度进一步提高了适体传感器的性能。在最佳条件下，计时库仑适体传感器的线性响应范围为 0.1-18.5 nM，检测限为 30 pM。结果表明，这种基于纳米粒子的放大策略为检测凝血酶提供了一种简单有效的方法。
