咖啡是世界三大饮料之一，其质量安全至关重要。其中，赭曲霉毒素A（OTA）是威胁咖啡质量安全的关键风险因子。2017年，国标《食品中真菌毒素限量》新增咖啡中OTA的限量标准，但是由于咖啡基质复杂，现有国标只提供仪器检测方法，而未提供快速检测方法。生物发光共振能量转移（BRET）是一种高灵敏的光学分析技术，其内源性发光特性，可减少基质背景荧光和散射光的干扰，对基质复杂样品的分析，具有明显优势。本项目拟以咖啡中的OTA为检测对象，纳米荧光素酶为BRET供体，近红外量子点为受体，纳米抗体为供/受体之间的连接臂，旨在探讨供/受体之间距离、供体量子产率、供体发射光谱与受体吸收光谱之间的重叠率以及免疫反应效率等因素，对BRET能量转移效率的影响，进而建立基于BRET的OTA高灵敏均相竞争免疫检测方法，并系统性开展抗咖啡基质干扰的研究。为建立抗复杂基质干扰，针对小分子物质的高灵敏快速检测方法提供理论依据。

Coffee is one of the three most-popular beverages in the world, and its safety are of paramount importance. Ochratoxin A (OTA) is a key risk factor threatening the safety of coffee. In order to minimize the health risk, the OTA limits in coffee has been established in 2017.Due to the complexity of the coffee matrix, the existing national standard only provides instrument detection methods in the absence of rapid detection methods. Bioluminescence Resonance Energy Transfer (BRET) is a novel highly sensitive optical analysis technique that eliminates the external light source, which can reduce the interference of matrix background and scattered light, has significant advantages for the analysis of complex matrix samples. This project intends to utilize OTA in coffee as the test object, nano-luciferase as BRET donor, near-infrared quantum dot as acceptor, and nanobody as the connecting arm between donor/receptor, aiming to explore the influence of the distance between donor and acceptor, donor quantum yield, overlap ratio between donor emission spectrum and acceptor absorption spectrum, and immune reaction efficiency on BRET energy transfer efficiency. To establish a novel method based on BRET homogeneous competitive immunoassay for sensitive dectection of OTA. Meanwhile, systematically research was conducted for anti-coffee matrix interference, to provide a theoretical basis for the establishment of anti-matrix interference, sensitivity and rapid detection methods for small molecules.