Zhang Shusheng, Zhong Hua, Ding Caifeng
Key Laboratory of Eco-chemical Engineering, Ministry of Education. College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, China.
Anal Chem. 2008 Oct 1;80(19):7206-12. doi: 10.1021/ac800847r. Epub 2008 Aug 30.
A novel and sensitive flow injection chemiluminescence assay for sequence-specific DNA detection based on signal amplification with nanoparticles (NPs) is reported in the present work. The "sandwich-type" DNA biosensor was fabricated with the thiol-functionalized capture DNA first immobilized on an Au electrode and hybridized with one end of target DNA, the other end of which was recognized with a signal DNA probe labeled with CuS NPs and Au NPs on the 3'- and 5'-terminus, respectively. The hybridization events were monitored by the CL intensity of luminol-H2O2-Cu(2+) after the cupric ions were dissolved from the hybrids. We demonstrated that the incorporation of Au NPs in this sensor design significantly enhanced the sensitivity and the selectivity because a single Au NP can be loaded with hundreds of signal DNA probe strands, which were modified with CuS NPs. The ratios of Au NPs, signal DNA probes, and CuS NPs modified on the gold electrode were approximately 1/101/103. A preconcentration process of cupric ions performed by anodic stripping voltammetry technology further increased the sensor performance. As a result of these two combined effects, this DNA sensor could detect as low as femtomolar target DNA and exhibited excellent selectivity against two-base mismatched DNA. Under the optimum conditions, the CL intensity was increased with the increase of the concentration of target DNA in the range of 2.0 x 10(-14)-2.0 x 10(-12) M. A detection limit of 4.8 x 10(-15) M target DNA was achieved.
本文报道了一种基于纳米颗粒（NPs）信号放大的新型灵敏流动注射化学发光法用于序列特异性DNA检测。“夹心型”DNA生物传感器的构建方法如下：首先将硫醇功能化的捕获DNA固定在金电极上，使其与目标DNA的一端杂交，目标DNA的另一端分别与3'-端标记有硫化铜纳米颗粒（CuS NPs）和5'-端标记有金纳米颗粒（Au NPs）的信号DNA探针识别。从杂交体中溶解出铜离子后，通过鲁米诺-H2O2-Cu(2+)的化学发光强度监测杂交事件。我们证明，在这种传感器设计中引入金纳米颗粒显著提高了灵敏度和选择性，因为单个金纳米颗粒可以负载数百条用硫化铜纳米颗粒修饰的信号DNA探针链。金电极上修饰的金纳米颗粒、信号DNA探针和硫化铜纳米颗粒的比例约为1/10/103。采用阳极溶出伏安法技术对铜离子进行预富集过程进一步提高了传感器性能。由于这两种效应的结合，这种DNA传感器能够检测低至飞摩尔浓度的目标DNA，并且对两碱基错配的DNA表现出优异的选择性。在最佳条件下，化学发光强度随目标DNA浓度在2.0×10(-14)-2.0×10(-12) M范围内的增加而增强。实现了对目标DNA的检测限为4.8×10(-15) M。
