基于“多色荧光-磁操纵-分子靶向”功能探针的多元真菌毒素荧光光谱/可视化双路径检测新方法研究

The principal objective of the program is to develop new aptasensing system for multiplex mycotoxins detection with a straightforward clean-up procedure for samples based on versatile nano-bioprobes. Firstly, this program presents a straightforward synthetic protocol of DNA1 bio-functionalized CdTe quantum dots (QDs) in a simple one-step reaction. One part of the DNA1 sequence controls the QDs passivation and serves as a ligand, while another part controls the biorecognition. Secondly, DNA2 bio-functionalized magnetic nanospheres will be fabricated as the biocarriers of the aptamer. Thirdly, versatile nanobioprobes with aptamer as a bridging linker, will be obtained based on the hybridization reaction between the DNA1, aptamer, and DNA2, respectively. When incubation of mycotoxins, the high specific binding between aptamers and targets will make some CdTe QDs fluorescent labels release from the magnetic nanospheres, which will be transferred by magnetic separation and measured by the fluorescence spectrometer. Meanwhile, the CdTe QDs fluorescent labels which are unreleased from the magnetic nanospheres will be accumulated by an external magnetic field for the visible sensor array fabrication. The dual fluorescent methods for simultaneous detection of multiplex mycotoxins based on versatile nano-bioprobes combine advanced optics and simple signal processing with low-cost, simplified procedure, and high efficiency, make the proposed high-throught simultaneous detection configuration attractive for food safety monitoring and other aptamer-based biomolecular recognition field.

本项目耦合量子点（QDs）荧光多通道性、磁性材料易操作性、适配体可设计和靶向识别性，先采用CdTe QDs生长过程中DNA1同步修饰法，制备不同粒径的DNA1-CdTe QDs；然后利用设计的适配体桥联所制备的DNA1-CdTe QDs与DNA2-磁性纳米球，研制“多色荧光-磁操纵-分子靶向”多功能纳米生物探针；通过探究不同磁球载体、界面功能化方法和杂交反应条件等因素，揭示纳米界面上生物分子之间的相互作用机制。进一步集合荧光光谱同时检测和荧光可视化阵列检测各自的技术优势，构建基于功能探针的多元真菌毒素双路径检测新方法；通过确立靶分子浓度与相应量子点荧光信号强度、光斑灰度/颜色亮度之间的定量关系，发展简便快捷、无需大型仪器、检测成本低的多元真菌毒素联合检测新技术；阐明多功能纳米生物探针在多靶标共性化分离富集与个性化选择检测方面的作用原理。因此项目的实施，具有较高的学术理论价值和应用前景。

农产品在其种植、生长、收割、储运及加工过程中都易滋生或接触到产毒真菌，从而导致真菌毒素进入食物链，直接促发胆管、食管、胃、肠、肺、肾、肝等脏器肿瘤的发生。目前，农产品中真菌毒素的检测技术存在灵敏度低、检测范围窄、耗时、重复性差等缺点，因此，开发高灵敏和高特异性的真菌毒素检测方法成为一项重大而紧迫的任务。本项目耦合量子点（QDs）荧光多通道性、磁性材料易操作性、适配体可设计和靶向识别性，利用末端带有功能基团的真菌毒素的适配体桥联作用制备“多色荧光-磁操纵-分子靶向”多功能纳米生物探针；通过探究不同磁球载体、界面功能化方法和杂交反应条件等因素，整合其他信号放大技术，建立了基于信号增强新型多功能探针的信号放大策略；发展了一系列基于新型纳米生物探针的典型真菌毒素（如赭曲霉毒素A（OTA）、伏马毒素B1（FB1））高灵敏检测新方法与新技术。研究表明，基于不同纳米粒子构建的磁控适配体比色、荧光、电化学传感体系能实现农产品（花生和玉米）中真菌毒素的快速、灵敏、同时检测，也为其它农产品的真菌毒素检测提供借鉴。由于纳米材料的吸附、信号放大、催化以及特殊的光电性能与增强光电信号的能力，显著地增强了真菌毒素传统检测方法的灵敏度。