Jiangsu Key Laboratory of New Power Batteries, Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, Jiangsu Key Laboratory of Biomedical Materials, National and Local Joint Engineering Research Center of Biomedical Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023, People's Republic of China.
Key Laboratory for Organic Electronics and Information Displays & Jiangsu Key Laboratory for Biosensors, Institute of Advanced Materials, Jiangsu National Synergetic Innovation Center for Advanced Materials (SICAM), Nanjing University of Posts and Telecommunications, Nanjing 210023, People's Republic of China.
Anal Chem. 2020 Apr 7;92(7):5302-5310. doi: 10.1021/acs.analchem.9b05849. Epub 2020 Mar 17.
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 cancer-derived exosomes, indicating its potential use in cancer diagnostics.
微小 RNA（miRNAs）在癌细胞衍生的外泌体中是重要的癌症生物标志物。在此，构建了一种用于检测外泌体 microRNA-122（miR-122）的灵敏杂交链式反应（HCR）电化学测定法。发夹 DNA（hpDNA）探针首先固定在金电极表面。在存在 miR-122 的情况下，hpDNA 的发夹结构可以打开，并通过两个辅助 DNA 发夹的交叉开口和杂交触发 HCR。从 HCR 产生的长缺口双链体用于捕获更多的 RuHex 并增加差分脉冲伏安法（DPV）的信号。在该测定中，通过同时固定 hpDNA 和短 12 个核苷酸单链 DNA（S-12）来精确控制金电极表面上的 hpDNA 探针的密度，提供了非常高的扩增效率。更重要的是，通过在引入靶标 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，该测定法具有非常高的灵敏度，检测限为毫微微摩尔级，线性范围为 9 个数量级，并且具有单错配区分的特异性。这种灵敏的电化学测定法可以成功评估来自不同癌症的外泌体中的 miR-122 浓度，表明其在癌症诊断中的潜在用途。
