基于特异银放大效应的界面分离抗干扰癌细胞检测体系研究

To avoid the interference effect of cells on the detection signal of traditional voltammetric biosensors, it is necessary to design novel biosensor configurations and develop new detection approaches. In the present proposal, a novel biosensor configuration has been designed, where the antibody will be immobilized on some other substrate surface for the recognition of cells, rather than on the electrode surface which is used for signal transformation. The configuration will be combined with gold nanoparticle-catalyzed silver enhancement reaction for signal generation and amplification. Then, the reduced silver will be detected by a bare electrode through the stripping voltammetry to further determine the amount of cells. As a result, the accuracy will be ameliorated through avoiding the interference effect of cells on detection signal and the sensitivity will be improved due to the signal amplification with silver enhancement reduction. Thus, an accurate and ultrasensitive electrochemical biosensor will be achieved for the detection of cancer cells and tissues. In the silver enhancement reaction, gold nanoparticle catalyst and hydrazine reductant will be combined together to avoid unspecific deposition of silver. Therefore, the accuracy will be further improved. This proposal is very important for the investigation of the biomolecular interaction and the application of catalysis of nanomaterials in the biodetection. The related research results may lead to an innovation in the electrochemical biosensor for cell detection, accelerate the application of electrochemcial biosensor for clinical application, and provide a new approach for cancer diagnosis. Furthermore, it is significative and valuable for reduction of the death rate of cancer disease in our country and the whole world.

为解决传统伏安型生物传感器中细胞对检测信号的干扰问题，提出新的细胞传感器构架及检测手段是必要的。本项目提出了分离细胞识别基体和信号转换电极的检测界面模式，即将细胞的识别抗体固定于电极以外的其他基体上，并结合银放大反应的信号产生方式，将对细胞的检测转化为对银离子的检测，最后用裸电极输出信号。这样既消除了细胞本身对检测的干扰，提高准确性；又通过银放大反应将放大信号，提高灵敏度，最终构建准确、超灵敏的电化学细胞传感器，对癌细胞和癌变组织进行检测。其中，银放大反应中纳米金催化剂和水合肼还原剂的结合，消除了银的非特异性沉积，进一步提高检测的准确性。该项目对于探讨生物分子间的相互作用和纳米催化作用在生物检测中的应用具有十分重要的意义。研究成果可能带来一次电化学细胞传感器构架的革新，加快其临床应用进程，为癌症诊断提供新的方法和依据，对降低我国以及全世界的癌症死亡率有重要的实际意义和科学研究价值。

癌症是威胁人类健康的头号杀手，有效的早期诊断是成功治疗癌症的关键所在。由于癌症标志物种类繁多，而且癌症早期时其浓度较低，难以准确检测。为此，本项目根据不同类型标志物的特点，将纳米材料和DNA扩增反应相结合，构建级联信号放大策略，设计了多种癌症标志物的检测方案，实现了痕量癌症标志物的超灵敏电化学检测。其中，本项目所报道的以DNA滚环扩增产物为模板合成的铜纳米颗粒为信号指示物的方案，与其它已报道的电化学检测方法相比，成功将前列腺特异抗原的检测限降低了4个数量级，并成功实现了人血清中前列腺特异抗原的可靠检测。在此基础上，本项目进一步提出并实现了分离细胞识别基底和信号转换电极的癌细胞检测界面模式，结合纳米材料-DNA信号放大网络，成功实现了目标癌细胞的可视化识别和超灵敏、高准确性的电化学检测。本项目在研究过程中提出了癌症标志物和癌细胞电化学检测的多种方案，对于探讨纳米材料在生物检测中的应用，开发生物化学检测的信号放大策略，研究癌细胞表面生物分子间的相互作用机制，促进癌症标志物和癌细胞检测新方法的开发有着十分重要的意义，为癌症的早期诊断、治疗、及药物输送等方面的研究提供了新的方法和依据，对降低我国以及全世界的癌症死亡率有重要的实际意义和科学研究价值。目前本项目已发表SCI收录的论文4篇，其中影响因子大于4.0的论文3篇；出版专著1章；申请专利2件。