同步检测多类兽药残留的流式荧光免疫分析方法研究

Food contamination caused by veterinary drugs is a world-wide public health concern. Multi-analyte immunoassay for simultaneously detecting multiple veterinary drugs in foods is in urgent need to solve the food safety issues in China. Traditional immunoassays usually focus on detecting single analyte or multiple analytes with similar chemical structures. Compared with parallel single-analyte immunoassay methods, the multi-analyte immunoassay offers high sample throughput, and could improve assay efficiency. The flow cytometry based multiplexed assay is an inherently and potentially promising analytical method to solve the bottleneck of multiple veterinary drug residue analysis. In the project we proposed, the multi-analyte profiling technology based multiple color fluorescence-encoded microsphere suspension array could be developed for simultaneously detecting multiple veterinary drug residues in food of animal origin. On the base of fluorescence-encoded microspheres, a new flow cytometry based multiplexed assay with traditional fluorescent dyes and novel fluorescent nanoparticles will also be developed for quantitatively determination of sulfonamides, quinolones, macrolides and lincosamides residues in foods of animal origin. After combining flow cytometry technology and fluorescent immunoassay, the flow cytometry based multiplexed assay could enrich the existing immunological analytical methods for chemical contaminants residues in foods of animal origin. The implement of the project would provide useful information for designing and improving new immunoassays for multiple chemical contaminates residue analysis.

兽药是重要的危害食品安全的化学污染物，动物性食品中的兽药多残留检测一直是食品安全检测领域的热点和难点。针对传统免疫分析方法难以同步定量检测多种不同种类兽药残留的技术瓶颈，本项目拟通过流式荧光免疫分析技术作为突破口，解决同时检测四种不同种类兽药多残留检测难题。本研究拟以荧光编码微球做免疫分析载体，利用不同种类荧光材料标记兽药抗原、抗体作为荧光检测探针，优化配对荧光编码微球与荧光标记物组合，在液相流体反应环境中进行多种兽药小分子抗原、抗体的竞争性免疫反应，建立多色流式荧光免疫分析方法，通过流式细胞仪同步定量分析特定荧光编码微球载体上荧光探针的荧光强度，同步定量检测动物性食品中磺胺类、喹诺酮类、大环内酯类和林可酰胺类等四类兽药残留。本研究可丰富现有的化学污染物多残留免疫检测技术，为动物性食品中兽药多残留免疫分析方法的设计、改进和发展提供必要的指导信息和新思路，具有重要理论意义和应用价值。

兽药和真菌毒素是重要的危害食品安全的化学污染物，动物性食品中的兽药和真菌毒素多残留检测一直是食品安全检测领域的热点和难点。针对传统免疫分析方法难以同步定量检测多种不同种类兽药与真菌毒素残留的技术瓶颈，本项目拟通过流式荧光免疫分析技术作为突破口，解决同时检测多种不同种类兽药与真菌毒素多残留检测难题。在前期工作基础上，项目团队基于四色荧光编码微球作为免疫分析载体，分别标记黄曲霉毒素、赭曲霉毒素、呕吐毒素和腐马菌素，以藻红蛋白作为标记信号探针，构建四色流式荧光免疫分析方法，同步检测牛奶中四种真菌毒素残留；基于多孔筛板作为免疫分析载体，以红色和绿色量子点作为标记探针，构建可视化荧光免疫分析方法，实现同步检测牛奶中可能存在的黄曲霉毒素和吡利霉素残留；此外，项目团队针对牛奶感染中常用的解热镇痛药氟尼辛，制备出同时识别氟尼辛和5-羟基氟尼辛小分子化合物的广谱特异性抗体，构建牛奶中氟尼辛和5-羟基氟尼辛残留的ELISA方法；针对林可酰胺类抗生素，制备同时识别林可霉素、克林霉素的广谱特异性抗体，并建立检测牛肉等动物源性食品中林可霉素和克林霉素残留的ELISA方法。截止2019年12月，项目团队在Journal of Agricultural and Food Chemistry（第一标注）、Meat Science（第一标注）、Food Analytical Methods（第一标注）、Food and Agricultural Immunology（第一标注）、Journal of Chromatography B（第二标注）等SCI收录期刊共计发表论文5篇，申请国家发明专利4项，培养硕士研究生1人。本项研究项目的成果丰富了现有的兽药与真菌毒素等化学污染物多残留免疫检测技术，为动物性食品中兽药与真菌毒素多残留免疫分析方法的设计、改进和发展提供必要的指导信息和新思路，具有重要理论意义和应用价值。