基于掺杂型量子点的时间分辨FRET体系的构建及生物传感

Time-resolved fluorescence resonance energy transfer (TR-FRET) combined low-background characteristic of time-resolved technique with homogeneous technique of fluorescence resonance energy transfer, which has more accurate results, more convenient experimental mode and higher sensitivity. The fluorophore probe is the key of the technique, at present, the probes are mainly lanthanide chelates owing to their long fluorescence life time (as long as 20~1000 μs). However, the luminescence of the lanthanide chelates is weak due to low luminescence quantum yields and small molar extinction coefficients. Also, photobleaching is serious for lanthanide chelates probes. The project is aimed at the key problem, the work would be performed as follows: Design and preparation of one series of long fluorescence life time（corresponding with lanthanide chelates）, strong photoluminescence, good stability and well water-dispersion doped Zinc-quantum dots，which would act as TR-FRET energy donor substituteded with lanthanide chelates, and Cadmium-quantum dots or fluorescent dyes act as energy acceptor, delaying measuring time, after short-lived background fluorescence disappeared, long-lived fluorescent complexes are determined to establish TR-FRET system and biosensor. It would be a powerful tool for biochemical and clinical detection such as antigen、 antibody and disease marker.

时间分辨荧光共振能量转移（TR-FRET）把时间分辨技术低背景特点和荧光共振能量转移均相实验方式融合在一起，使它具有更准确的实验结果、更灵活的实验方式、更高的灵敏度等优势。荧光标记物是该技术的关键所在，目前普遍使用的稀土配合物存在着较低的荧光量子产率、较小的摩尔吸光系数及光漂白的问题，降低了TR-FRET的灵敏度和稳定性。本项目围绕这一关键问题，拟通过设计和制备长荧光寿命（与稀土配合物相当）、高荧光强度和良好光稳定性的掺杂型锌系量子点；从而替代稀土配合物作为TR-FRET能量供体，镉系量子点或荧光染料作为能量受体，通过延迟测量时间，待本底短寿命荧光消失之后, 才对能量转移的长寿命荧光进行测定，从而构建TR-FRET体系及其生物传感研究，可望成为抗原、抗体、疾病标志物等生命分析强有力的工具。

时间分辨荧光共振能量转移（TR-FRET）生物传感器的研究处于起步阶段。在构建高灵敏TR-FRET生物传感器的过程中尚存在一系列问题，如目前普遍使用的荧光标记物-稀土配合物荧光量子产率低、摩尔吸光系数小及光漂白；分析对像窄、检测灵敏度低等。本项目围绕这些关键问题，设计和制备了不同形态和尺寸的掺杂型锌系荧光量子点（Mn:ZnS,Mn:ZnSe, Mn:ZnS/ZnO, Mn:ZnSe/ZnS等），它们具有长荧光寿命（与稀土配合物相当）、高荧光强度和良好光稳定性等特点，从而替代稀土配合物作为TR-FRET 能量供体；制备作为TR-FRET能量受体的纳米金胶，并对这些材料进行有利于抗原、抗体等生物分子相结合的功能化修饰；通过采用时间分辨荧光分析法，成功地构建了一系列针对血清中药物分子（硫普罗宁（tiopronin）、吲达帕胺（indapamide））蛋白、疾病标志物（血管内皮生长因子(VEGF)、血小板衍生因子（PDGF））的生物传感器，可望成为抗原、抗体、疾病标志物等生命分析强有力的工具。