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 210097 , P. R. China.
Key Laboratory for Organic Electronics and Information Displays & Institute of Advanced Materials, Jiangsu National Synergetic Innovation Center for Advanced Materials (SICAM)) , Nanjing University of Posts & Telecommunications , Nanjing 210023 , P. R. China.
Anal Chem. 2018 Apr 3;90(7):4507-4513. doi: 10.1021/acs.analchem.7b04863. Epub 2018 Mar 15.
Sensitive and specific detection of tumor exosomes is of great significance for early cancer diagnosis. In this paper, we report an aptamer strategy for exosome detection based on aptamer recognition-induced multi-DNA release and cyclic enzymatic amplification. First, we use aptamer-magnetic bead bioconjugates to capture tumor exosomes derived from LNCaP cells, leading to the release of three kinds of messenger DNAs (mDNAs). After magnetic separation, the released mDNAs hybridized with the probe DNAs immobilized on a gold electrode. Electroactive Ru(NH) was used as the signal reporter because of its electrostatic attraction to DNA. Subsequent Exo III cyclic digestion caused the electrochemical signal to "turn off". Because the electrochemical signal reflects the concentration of Ru(NH) and the concentration of Ru(NH) is correlated with the mDNA concentration, which is correlated with the exosome concentration, the tumor exosomes can be detected by examining the decrease in the peak current of Ru(NH). In this paper, the signal was amplified by the numerous mDNAs released from the magnetic bead and the Exo III-assisted mDNA recycling. Under the optimal conditions, a detection limit down to 70 particles/μL was achieved, which is lower than the LODs of most currently available methods. Furthermore, this assay can be used to detect tumor exosomes in complex biological samples, demonstrating potential application in real sample diagnosis.
敏感而特异地检测肿瘤外泌体对于早期癌症诊断具有重要意义。本文报道了一种基于适配体识别诱导的多 DNA 释放和循环酶促扩增的外泌体检测适配体策略。首先，我们使用适配体-磁珠偶联物捕获源自 LNCaP 细胞的肿瘤外泌体，导致三种信使 DNA（mDNA）的释放。在磁分离后，释放的 mDNA 与固定在金电极上的探针 DNA 杂交。由于其与 DNA 的静电吸引力，电化学活性 Ru(NH)被用作信号报告分子。随后，Exo III 循环消化导致电化学信号“关闭”。由于电化学信号反映了 Ru(NH)的浓度，而 Ru(NH)的浓度与 mDNA 浓度相关，mDNA 浓度与外泌体浓度相关，因此可以通过检测 Ru(NH)峰电流的减少来检测肿瘤外泌体。在本文中，通过从磁珠上释放的大量 mDNA 和 Exo III 辅助的 mDNA 循环来放大信号。在最佳条件下，检测限低至 70 个颗粒/μL，低于大多数现有方法的 LODs。此外，该测定法可用于检测复杂生物样品中的肿瘤外泌体，表明其在实际样品诊断中有潜在的应用。
