Electrochemical Sensing of Exosomal MicroRNA Based on Hybridization Chain Reaction Signal Amplification with Reduced False-Positive Signals
Electrochemical Sensing of Exosomal MicroRNA Based on Hybridization Chain Reaction Signal Amplification with Reduced False-Positive Signals
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DOI：
10.1021/acs.analchem.9b05849
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发表时间：
2020-04-07
影响因子：
7.4
通讯作者：
Shen, Qingming
中科院分区：
文献类型：
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作者：
Guo, Qunqun;Yu, Yongqi;Shen, Qingming
癌细胞来源的外泌体中的微小RNA（miRNAs）是重要的癌症生物标志物。本文中，制备了用于检测外泌体microRNA-122（miR-122）的灵敏的杂交链反应（HCR）电化学测定。首先将发夹DNA（hpDNA）探针固定在金电极表面。在miR-122的存在下，hpDNA的发夹结构可以被打开，并通过两个辅助DNA发夹的交叉打开和杂交来触发HCR。由HCR产生的长切口双螺旋用于捕获更多的RuHex并增加差示脉冲伏安法（DPV）的信号。在该测定中，hpDNA探针在金电极表面上的密度通过hpDNA和短的12个核苷酸的单链DNA（S-12）的同时固定而精确控制，提供了非常高的扩增效率。更重要的是，通过在引入靶miR-122之前应用核酸外切酶I（Exo I）可以减少或完全消除假阳性信号。在最佳条件下，该测定提供了非常高的灵敏度，具有阿托摩尔水平的检测限，线性范围为9个数量级，并在单一错配歧视的特异性。这种灵敏的电化学检测方法可以成功地评估不同癌症来源的外泌体中miR-122的浓度，表明其在癌症诊断中的潜在用途。
MicroRNAs (miRNAs) in cancer cell-derived exosomes are important cancer biomarkers. Herein, a sensitive hybridization chain reaction (HCR) electrochemical assay was fabricated for the detection of exosomal microRNA-122 (miR-122). The hairpin DNA (hpDNA) probes were first immobilized on the surface of a gold electrode. In the presence of miR-122, the hairpin structure of the hpDNA could be opened and triggered the HCR through the cross-opening and hybridization of two helper DNA hairpins. Long nicked double helixes generated from HCR are used to capture more RuHex and increase the signal of differential pulse voltammetry (DPV). In this assay, the density of the hpDNA probes on the surface of the gold electrode was precisely controlled by the simultaneous immobilization of hpDNA and short 12 nucleotides single-stranded DNA (S-12), providing a very high amplification efficiency. More importantly, the false positive signal could be reduced or completely eliminated by applying exonuclease I (Exo I) before the introduction of target miR-122. Under optimal conditions, the assay offers very high sensitivity with an attomolar level detection limit, a linear range with 9 orders of magnitude, and specificity in single mismatch discrimination. This sensitive electrochemical assay could successfully evaluate the miR-122 concentration in different cancerderived exosomes, indicating its potential use in cancer diagnostics.