无标记电化学 DNA 传感的单靶多触发杂交链式反应策略。
Label-free electrochemical DNA sensing with a one-target-multitriggered hybridization chain reaction strategy.
机构信息
State Key Laboratory of Analytical Chemistry for Life Science, Department of Chemistry, Nanjing University, Nanjing 210093, P.R. China.
出版信息
Analyst. 2013 Oct 21;138(20):5995-6000. doi: 10.1039/c3an01212c. Epub 2013 Aug 15.
A one-target-multitriggered hybridization chain reaction (MHCR) strategy was designed for ultrasensitive electrochemical detection of DNA by combining the isothermal strand-displacement polymerase reaction (ISDPR) with the DNA self-assembly on a DNA sensor surface. The sensor was constructed by immobilizing a hairpin-like capture probe (CP) on a gold electrode via an Au-S bond. The ISDPR was triggered by the hybridization of the target DNA to open the CP and primer to anneal the complementary part in the bottom of the exposed stem and the extension of the primer in the presence of dNTPs and polymerase. Each target copy could produce a few opened CPs. Afterwards, the other part of the exposed stem acted as an initiator to trigger the hybridization chain reaction (HCR) when incubated with two hairpin monomers. Using Ru(NH3)6 as an electrochemically active indicator to interact with the MHCR product, the amperometric response demonstrated a perfect multiple amplification effect. The constructed sensor showed a high sensitivity for detection of the target DNA in a linear range from 0.1 fM to 10 pM, a detection limit down to 0.02 fM (3σ) and good selectivity for base discrimination. This method did not need any modification or labelling process. The proposed strategy provides a powerful tool for cascade signal amplification and has a wide potential application in bioanalysis.
一种单靶多触发杂交链式反应(MHCR)策略通过将等温链置换聚合反应(ISDPR)与 DNA 传感器表面上的 DNA 自组装相结合,被设计用于超灵敏电化学检测 DNA。传感器通过金 - 硫(Au-S)键将发夹状捕获探针(CP)固定在金电极上。ISDPR 由目标 DNA 与 CP 杂交引发,打开 CP 并使引物与暴露茎底部的互补部分退火,在 dNTPs 和聚合酶存在下延伸引物。每个目标副本都可以产生几个打开的 CP。之后,当与两个发夹单体孵育时,暴露茎的另一部分充当引发剂来触发杂交链式反应(HCR)。使用 Ru(NH3)6 作为电化学活性指示剂与 MHCR 产物相互作用,安培响应显示出完美的多重扩增效果。所构建的传感器在 0.1 fM 至 10 pM 的线性范围内对目标 DNA 的检测具有高灵敏度,检测限低至 0.02 fM(3σ),并且对碱基具有良好的选择性。该方法不需要任何修饰或标记过程。该策略为级联信号放大提供了强大的工具,在生物分析中有广泛的潜在应用。