Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran; Student Research Committee, Baqiyatallah University of Medical Sciences, Tehran, Iran.
Department of Textile and Polymer Engineering, Yazd Branch, Islamic Azad University, Yazd, Iran.
Talanta. 2023 Jan 15;252:123863. doi: 10.1016/j.talanta.2022.123863. Epub 2022 Aug 24.
In this study, the dual signal-labeled hairpin-structured DNA (dhDNA)-based probes have been developed to construct a novel nano-biosensor. This one hairpin-structured probe consists of a thiolated methylene blue-labeled hairpin capture probe (MB-HCP) as an inner reference probe and a ferrocene-modified anti-miRNA-21 DNA probe (Fc-AP-21). This novel integrated structure of MB-HCP and Fc-AP-21 was designed on one sensing interface for sensitive and simultaneous detection of the miRNA-141 and miRNA-21 in one single assay. The proposed strategy has a good ability to reduce the interference of environmental factors and it was designed to control the initial responses of Fc-AP to MB-HCP ((I/I)) at a 1:1 ratio, which is desirable for further increase in the sensitivity and signal-to-noise ratio of the biosensor operation. Besides, the biosensor was first prepared by immobilizing the dhDNA (Fc-AP-21/MB-HCP) onto the modified glassy carbon electrode. After hybridization with the anti-miRNA-141 complementary sequence (ACP-141), the dhDNA structure was compelled to open and form the final structure of the biosensor. Also, the miRNA-141 and miRNA-21 dissociate duplex structures due to the highly matched sequences between the miRNA-141 and ACP-141 and the miRNA-21 and Fc-AP-21. A linear relationship was found between the logarithm of miRNA-141 and miRNA-21 concentrations and the signal changes. This feature was used to detect the two miRNAs. This sensitive biosensor provided low detection limits of 0.89 and 1.24 fM for the miRNA-141 and miRNA-21, respectively. Also, it has wide linear ranges of 2.0 to 10 fM, with highly selective and accurate results for its application in plasma samples. Therefore, this strategy can be promising as a suitable platform for simultaneous and early detection of various cancer biomarkers.
在这项研究中，开发了双信号标记发夹结构 DNA（dhDNA）探针来构建新型纳米生物传感器。该发夹结构探针由一个巯基化亚甲基蓝标记的发夹捕获探针（MB-HCP）作为内参探针和一个修饰有二茂铁的抗 miRNA-21 DNA 探针（Fc-AP-21）组成。这种新型的 MB-HCP 和 Fc-AP-21 集成结构设计在一个传感界面上，用于在单个测定中同时灵敏地检测 miRNA-141 和 miRNA-21。所提出的策略具有很好的减少环境因素干扰的能力，并设计为控制 Fc-AP 对 MB-HCP 的初始响应（I/I）在 1:1 的比例，这有利于进一步提高生物传感器操作的灵敏度和信噪比。此外，该生物传感器首先通过将 dhDNA（Fc-AP-21/MB-HCP）固定在修饰的玻碳电极上制备。与抗 miRNA-141 互补序列（ACP-141）杂交后，dhDNA 结构被迫打开并形成生物传感器的最终结构。同样，由于 miRNA-141 和 ACP-141 之间以及 miRNA-21 和 Fc-AP-21 之间高度匹配的序列，miRNA-141 和 miRNA-21 解离双链结构。发现 miRNA-141 和 miRNA-21 浓度与信号变化之间存在对数线性关系。该特性用于检测这两种 miRNA。该灵敏的生物传感器为 miRNA-141 和 miRNA-21 分别提供了 0.89 和 1.24 fM 的低检测限。此外，它具有 2.0 至 10 fM 的宽线性范围，对其在血浆样品中的应用具有高度选择性和准确性。因此，该策略有望成为同时和早期检测各种癌症生物标志物的合适平台。
