基于纳米金磁复合微粒的布鲁氏菌抗体定量免疫层析分型检测技术的研究

Brucellosis is a zoonotic disease caused by members of the genus Brucella．It is widely popular in the world，especially in developing countries．Brucella is a facultative intracellular parasitic bacterium．Due to its strong invasion and multiple approaches of transmission，patients show persistent infection，which is usually known as incurable disease．Infectious animal mainly caused abortion，which leads to serious loss for livestock industry．a simple, economical and rapid, high-sensitivity method of detection for early diagnosis of Brucellosis is critical for control and prevention of this disease. Previous research achievements, we report on a facile method to synthesize novel Fe3O4/Au/Fe3O4 nanoparticles (nanoflowers) that integrate hybrid components and surface types.We demonstrate that relative to conventional nanoparticles with core/shell configuration, such nanoflowers not only retain their surface plasmon property but also allow for 170% increase in the saturation magnetization and 23% increase in the conjugation efficiency due to the synergistic co-operation between the hierarchical structures. We established a method of colloidal gold immunochromatographic to classification diagnose the antibody of Brucella. immunochromatographic strip was prepared with Brucella BP26 protein as diagnosis antigen and OMP31 as classification diagnosis antigen. In this study，the lateral flow immunoassay utilizing novel Fe3O4/Au/Fe3O4 nanoparticles instead of colloidal gold, taking advantage of the magnetism, optics and immunochromatography, we will establish a rapid, high-sensitive and quantitative detection of Brucella.

布鲁氏菌病（简称布病）是由布鲁氏菌引起的一类动物源性人兽共患病，在世界范围内广泛流行，尤其是发展中国家。布鲁氏菌是一种兼性胞内寄生致病菌，侵袭力强、传染途径多，人感染后呈持续性感染，目前尚无法根治；家畜感染后主要引发母畜流产，给畜牧业生产造成严重损失。建立一种适用于基层的快速、高灵敏度、定量的即时检测技术是布鲁氏菌病早期诊断和疾病控制的关键。课题组首次合成出新型向日葵形貌的纳米金磁复合微粒，增加了核壳型纳米金磁微粒的饱和磁化强度170%和增加了抗体蛋白的偶联效率23%，且具有纳米金粒子的等离子共振吸收特性;获得了布鲁氏菌膜蛋白BP26、OMP31作为抗原建立了布鲁氏菌免疫层析分型检测技术。本研究将以新型纳米金磁微粒作为标记材料，以BP26为诊断抗原，OMP31作为分型诊断抗原建立一种对布鲁氏菌病的简单快速、高灵敏、定量分型的检测技术。

布鲁氏菌病（简称布病）是由布鲁氏菌引起的一类动物源性人兽共患病，在世界范围内广泛流行，尤其是发展中国家。布鲁氏菌是一种兼性胞内寄生致病菌，侵袭力强、传染途径多，人感染后呈持续性感染，目前尚无法根治；家畜感染后主要引发母畜流产，给畜牧业生产造成严重损失。建立一种适用于基层的快速、高灵敏度、定量的即时检测方法是布鲁氏菌病早期诊断和疾病控制的关键。本研究通过原核表达技术获得了布鲁氏菌的BP26蛋白、OMP31蛋白、VirB5蛋白、SP41蛋白、PGM蛋白和分离纯化了脂多糖成分，通过ELISA、WB及其胶体金免疫渗滤技术和免疫层析技术系统验证了作为检测抗体的抗原性，结果发现： BP26能够诊断抗原，OMP31作为分型诊断抗原外，还发现布鲁氏菌的脂多糖成分能够应用布鲁氏菌抗体的检测。本项目合成了出新型向日葵形貌的纳米金磁复合微粒，增加了核壳型纳米金磁微粒的饱和磁化强度170%和增加了抗体蛋白的偶联效率23%，且具有纳米金粒子的等离子共振吸收特性;将获得的布鲁氏菌膜蛋白BP26、OMP31作为抗原建立了布鲁氏菌免疫层析分型检测技术。建立了纳米金磁复合微粒的制备方法以此为标记材料建立了具有方便快速、高灵敏、现场能够快速、定量、分型检测布鲁氏菌病抗体的免疫层析检测系统。通过本研究还研制了的布鲁氏菌病抗体快速检测渗滤卡和试纸卡并获得授权专利，与公司企业合作形成产品，进行了推广应用。本研究项目发表研究论文3篇、获得授权专利1项，培养硕士研究生3人、本科毕业生6人。