García T, Casero E, Revenga-Parra M, Martín-Benito J, Pariente F, Vázquez L, Lorenzo E
Departamento de Química Analítica y Análisis Instrumental, Universidad Autónoma de Madrid, Campus de Cantoblanco, 28049 Madrid, Spain.
Biosens Bioelectron. 2008 Oct 15;24(2):184-90. doi: 10.1016/j.bios.2008.03.034. Epub 2008 Apr 4.
The preparation of DNA-sensing architectures based on gold nanoparticles (Au-NPs) in conjunction with an "in situ" prepared ruthenium complex as a new route to improve the analytical properties of genosensors is described. In the development of these architectures several strategies to obtain Au-NPs modified gold electrodes (Au-NP/Au) have been essayed, in particular covalent binding and electrochemical deposition from a solution containing Au-NPs previously synthesized. UV-vis absorption measurements in conjunction with transmission electron microscope (TEM) images reveal that the synthesized Au-NPs are stable for at least 4 weeks and have a narrow size distribution. Atomic force microscopy (AFM) was employed to characterize the morphology and to estimate the Au-NPs surface coverage of the modified gold electrodes obtained following the different modification strategies. In order to assess the utility of these architectures as DNA-sensing devices, a thiolated capture probe sequence from Helicobacter pylori was immobilized onto the as-prepared surface. This sequence was chosen as a case of study within the framework of developing approaches of wide applicability. The hybridization event is detected using a water-soluble pentaamin ruthenium [3-(2-phenanthren-9-yl-vinyl)-pyridine] complex (Ru(NH(3))(5)L) prepared "in situ". This complex, due to its intercalative character, is able to bind to double stranded DNA more efficiently than to single stranded DNA. In addition, the metal provides with a redox center that can be used as an electrochemical indicator. On the basis of this strategy, complementary target sequences of H. pylori have been detected over the range of 40-800 pmol with a detection limit of 25+/-2 pmol.
本文描述了基于金纳米颗粒(Au-NPs)并结合“原位”制备的钌配合物来制备DNA传感结构,这是一种改善基因传感器分析性能的新途径。在这些结构的开发过程中,尝试了几种获得Au-NPs修饰金电极(Au-NP/Au)的策略,特别是共价结合以及从含有先前合成的Au-NPs的溶液中进行电化学沉积。紫外可见吸收测量结合透射电子显微镜(TEM)图像表明,合成的Au-NPs至少在4周内是稳定的,并且尺寸分布狭窄。采用原子力显微镜(AFM)来表征形态并估计通过不同修饰策略获得的修饰金电极上Au-NPs的表面覆盖率。为了评估这些结构作为DNA传感装置的实用性,将来自幽门螺杆菌的硫醇化捕获探针序列固定在制备好的表面上。在开发具有广泛适用性的方法框架内,选择该序列作为一个研究案例。使用“原位”制备的水溶性五氨基钌[3-(2-菲-9-基-乙烯基)-吡啶]配合物(Ru(NH(3))(5)L)检测杂交事件。由于其嵌入特性,该配合物与双链DNA的结合效率高于与单链DNA的结合效率。此外,该金属提供了一个可作为电化学指示剂的氧化还原中心。基于此策略,已在40 - 800 pmol范围内检测到幽门螺杆菌的互补靶序列,检测限为25±2 pmol。