基于“开关”型FRET近红外比率荧光探针的构建及信号放大技术用于乙肝病毒DNA的循环检测与成像研究

In recent years, the incidence of hepatitis B has been rising. Because of its highly contagious nature, it poses a serious threat to human life and property. At present, there are many deficiencies in the analysis of hepatitis B virus at home and abroad. Thus, establishing an analytical strategy which has the advantages of convenience, quickness, strong sensitivity, high selectivity is looming ahead. The innovations of this project are synthesizing N, S and F co-doped graphene quantum dots modified by ionic liquid for the first time, at the same time, innovatively synthetizing gold nanoparticles with NIR fluorescence and labeling DNA with blue-emitting doped graphene quantum dots. What’s more, it’s the first time the novel and smart dual-emitting FRET system with quantum dots as energy donor and NIR-emitting gold nanoparticles as energy acceptor is established. Based on the built FRET system, a technology based on a ‘switch’ type FRET ratio method and enzyme signal cyclic amplification is used to perform highly sensitive and selective analysis of the hepatitis B virus DNA and explore their potential value in cell and vivo imaging. The research results of this project are conducive to the development of new strategies for the synthesis and application of multifunctional nanoparticles, which will be a useful attempt to further promote the new developments in the field of biochemistry and biomedicine, and provide scientific reference for the early diagnosis and treatment of major diseases.

近年来，乙型肝炎发病一直呈上升趋势，由于其具有极强的传染性，给人类的生命财产带来了严重威胁。目前国内外乙肝病毒的分析方法均存在其缺陷，因此建立一种简便快捷、高灵敏度、高选择性的乙肝病毒DNA分析方法迫在眉睫。本项目的创新之处在于，首次合成了具有蓝色荧光的N, S, F三元素共掺杂离子液体修饰的石墨烯量子点，同时创新性地合成了发射近红外荧光的金纳米粒子，并以掺杂石墨烯量子点标记DNA探针，构建了以掺杂石墨烯量子点为能量供体，近红外金纳米粒子为能量受体的新颖、巧妙的双发光FRET探针体系。利用“开关”型荧光信号转换比率法和DNA酶循环扩增信号放大技术对乙肝病毒DNA进行高灵敏度、高选择性分析，探究其在细胞及活体成像的潜在应用价值。本项目研究成果有利于多功能纳米粒子合成及应用新策略开发，并为进一步推动生物化学和生物医学领域的新发展作了有益尝试，为重大疾病的早期诊断和治疗提供科学参考。

按照计划，通过控制合成纳米材料的结构、形貌、尺寸及其它因素，揭示诸因素等对材料性能的影响规律，为材料的实践应用和性能改进提供了理论与实验借鉴。制备了具有特定物理、化学、光学和电学性质的系列纳米材料，并开展了其在不同领域的应用研究。主要包括系列合成的纳米材料在荧光传感领域的应用以及纳米复合材料增强对目标物的光电和电化学检测性能等。（1）研制了一种新型AuNPs-DNA混合传感器，用于荧光检测水溶液中的痕量汞(II)离子(Hg2+)。（2）研究了以金属-有机框架为载体的双金属掺杂比率荧光纳米材料作为生物血清胆固醇定量测定的高效探针以及相关合成、表征、机理和DFT计算。（3）系统探索了Eu(III)和Zr(IV)杂化的双发射双金属有机框架用于油炸及烘焙食品中微量丙烯酰胺的比率荧光传感。（4）详尽探究了金属有机框架核酸适配体荧光传感器，用于酶催化扩增吸附DNA超灵敏测定ATP及其逻辑门研究。（5）探讨了基于GQDs的“On-Off-On”转换开关型荧光纳米传感器对Cu2+和硫普罗宁的快速和无标记双重检测。除了在荧光领域的探索外，还拓展了项目合成的系列纳米材料在电化学领域的应用范围。（1）探究了AuNPs/N,S,P-CQDs作为信号增强剂对环境内分泌干扰物BPA的高选择性电化学适配体传感及其电化学动力学研究。（2）探索了AuNPs/N,S-GQDs纳米复合材料的电化学行为和动力学研究，将其开发为一种新型高灵敏度和高选择性的无标记扩增BPA适配体传感器。（3）探讨了基于MIP和B,N,F-CQDs/AgNPs的新型电化学传感器，用于塑料制品中高选择性超痕量BPS增强特异性识别和双信号放大的实验和理论研究。（4）研究了Ag@Cu2O和CdS/g-C3N4/ITO纳米复合异质结材料，通过抗原抗体形成三明治夹心结构，实现对癌胚抗原CEA的特异性识别和多重信号放大检测。