State Key Laboratory of Analytical Chemistry for Life Science and Collaborative Innovation Center of Chemistry for Life Sciences, School of Chemistry and Chemical Engineering, Nanjing University , Nanjing 210023, China.
ACS Appl Mater Interfaces. 2017 Sep 27;9(38):33360-33367. doi: 10.1021/acsami.7b11524. Epub 2017 Sep 14.
In this study, we proposed a nanopore-based electrochemiluminescence (ECL) sensor combined with duplex-specific nuclease (DSN)-assisted target recycling amplification to detect microRNAs. Because of the synergetic effect of electrostatic repulsion and volume exclusion of gold nanoparticle-labeled DNA capture (DNA-Au NPs) to the negatively charged luminol anion probe, the DNA-Au NP-modified anodized aluminum oxide (AAO) nanopore electrode exhibited high ECL decline in comparison with the bare AAO electrode. Upon the introduction of DSN and target microRNA, the specific DNA-RNA binding and enzyme cleaving could trigger the detachment of capture DNA from the membrane surface, resulting in uncapping of AAO and an increased ECL signal. For comparison, positively charged Ru(bpy) was used as the ECL probe instead of luminol. Because the electrostatic attraction effect between DNA and Ru(bpy) is partially offset by the volume exclusion effect of Au NPs, the AAO electrode modified with only DNA capture is more suitable for the Ru(bpy) case. In our experiment, the case of negatively charged luminol combined with the synergetic effect of electrostatic repulsion and volume exclusion of DNA-Au NPs provides a quantitative readout proportional to the target microRNA concentration in the range of 1.0 fM to 1.0 nM, with a lower detection limit of 1 fM.
在本研究中，我们提出了一种基于纳米孔的电化学发光（ECL）传感器，结合双链特异性核酸酶（DSN）辅助的靶标循环扩增来检测 microRNA。由于金纳米粒子标记的 DNA 捕获（DNA-Au NPs）对带负电荷的鲁米诺阴离子探针的静电排斥和体积排除的协同作用，与裸 AAO 电极相比，DNA-Au NP 修饰的氧化铝纳米孔电极表现出更高的 ECL 衰减。引入 DSN 和靶标 microRNA 后，特异性的 DNA-RNA 结合和酶切割会触发捕获 DNA 从膜表面脱离，导致 AAO 去帽和 ECL 信号增加。相比之下，带正电荷的 Ru(bpy) 被用作 ECL 探针而不是鲁米诺。由于 DNA 和 Ru(bpy) 之间的静电吸引作用部分被 Au NPs 的体积排除效应抵消，因此仅修饰 DNA 捕获的 AAO 电极更适合 Ru(bpy) 情况。在我们的实验中，带负电荷的鲁米诺与 DNA-Au NPs 的静电排斥和体积排除的协同作用相结合的情况提供了与靶标 microRNA 浓度成正比的定量读数，其在 1.0 fM 至 1.0 nM 的范围内的检测下限为 1 fM。
