Lou Jing, Liu Shanshan, Tu Wenwen, Dai Zhihui
Jiangsu Collaborative Innovation Center of Biomedical Functional Materials and Jiangsu Key Laboratory of Biofunctional Materials, College of Chemistry and Materials Science, Nanjing Normal University , Nanjing, 210023, P. R. China.
Anal Chem. 2015 Jan 20;87(2):1145-51. doi: 10.1021/ac5037318. Epub 2015 Jan 5.
A novel strategy for highly sensitive electrochemiluminescence (ECL) detection of DNA was proposed based on site-specific cleavage of BamHI endonuclease combined with the excellent ECL activity of graphene quantum dots (GQDs) and bidentate chelation of the dithiocarbamate DNA (DTC-DNA) probe assembly. The difference between photoluminescence and ECL spectral peaks suggested that a negligible defect existed on the GQDs surface for generation of an ECL signal. The formed DTC-DNA was directly attached to the gold surface by bidentate anchoring (S-Au-S bonds), which conferred a strong affinity between the ligands and the gold surface, increasing the robustness of DNA immobilization on the gold surface. BamHI endonuclease site-specifically recognized and cleaved the duplex symmetrical sequence, which made the double-stranded DNA fragments and GQDs break off from the electrode surface, inducing a decrease of the ECL signal. Using hepatitis C virus-1b genotype complementary DNA (HCV-1b cDNA) as a model, a novel signal-off ECL DNA biosensor was developed based on variation of the ECL intensity before and after digestion of the DNA hybrid. Electrochemical impedance spectroscopy confirmed the successful fabrication of the ECL DNA biosensor. This ECL biosensor for HCV-1b cDNA determination exhibited a linear range from 5 fM to 100 pM with a detection limit of 0.45 fM at a signal-to-noise ratio of 3 and showed satisfactory selectivity and good stability, which validated the feasibility of the designed strategy. The proposed strategy may be conveniently combined with other specific biological recognition events for expansion of the biosensing application, especially in clinical diagnoses.
基于BamHI核酸内切酶的位点特异性切割，结合石墨烯量子点（GQDs）优异的电化学发光（ECL）活性和二硫代氨基甲酸盐DNA（DTC-DNA）探针组装体的双齿螯合作用，提出了一种用于DNA高灵敏度电化学发光检测的新策略。光致发光和ECL光谱峰之间的差异表明，GQDs表面存在可忽略不计的缺陷以产生ECL信号。形成的DTC-DNA通过双齿锚定（S-Au-S键）直接附着在金表面，这赋予了配体与金表面之间的强亲和力，增加了DNA固定在金表面的稳定性。BamHI核酸内切酶位点特异性识别并切割双链对称序列，使双链DNA片段和GQDs从电极表面脱落，导致ECL信号降低。以丙型肝炎病毒1b基因型互补DNA（HCV-1b cDNA）为模型，基于DNA杂交消化前后ECL强度的变化，开发了一种新型的信号关闭型ECL DNA生物传感器。电化学阻抗谱证实了ECL DNA生物传感器的成功制备。这种用于测定HCV-1b cDNA的ECL生物传感器在5 fM至100 pM范围内呈线性，在信噪比为3时检测限为0.45 fM，具有令人满意的选择性和良好的稳定性，验证了所设计策略的可行性。所提出的策略可以方便地与其他特定的生物识别事件相结合，以扩展生物传感应用，特别是在临床诊断中。
