IgG-Biotin-Avidin-HRP信号转导探针结合免疫比色法快速检测克罗诺阪崎肠杆菌  

Rapid Detection of Cronobacter sakazakii by IgG-Biotin-Avidin-HRP Combined with Colorimetric Immunoassay

作　　者：叶玲娴 黄浩然 柳露 吴遵义 黄建锋 赵广英 窦文超 Ye Lingxian;Huang Haoran;Liu Lu;Wu Zunyi;Huang Jianfeng;Zhao Guangying;Dou Wenchao(Food Safety Key Laboratory of Zhejiang Province,College of Food Science and Biotechnology,Zhejiang Gongshang University,Hangzhou 310018;Zhejiang Fangyuan Test Group Co.,Ltd.,Hangzhou 310018)

机构地区：[1]浙江工商大学食品与生物工程学院,浙江省食品安全重点实验室,杭州310018 [2]浙江省方圆检测集团股份有限公司,杭州310018

出　　处：《中国食品学报》2020年第7期237-243,共7页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家自然科学基金项目(30571623);浙江省自然科学基金项目(LY17C200003);浙江省公共创新平台分析测试项目(2018C37056);浙江省新苗人才计划项目(2017R408054,2017R408053)。

摘　　要：本文建立了一种快速检测克罗诺阪崎肠杆菌的新型免疫比色法。用抗体修饰的磁球(MNPs-IgG)作为捕获探针分离、富集克罗诺阪崎肠杆菌,再通过免疫夹心法分别结合生物素标记的克罗诺阪崎肠杆菌抗体(Biotin-IgG)和亲和素标记的辣根过氧化物酶(Avidin-HRP),形成的复合物与底物四甲基联苯胺(3,3′,5,5′-Tetramethylbenzidine,TMB)发生显色反应,经硫酸终止后,有色产物的吸光度值与克罗诺阪崎肠杆菌浓度的对数有良好的线性关系,在103~107CFU/mL范围内,线性方程为y=0.2353x-0.2307。本研究成功设计优化了基于比色法的克罗诺阪崎肠杆菌定量检测新方法,检测限为8.68×10^2CFU/mL(S/N=3),加标样品检出率96%,检测时间不超过2.5 h,为实际应用与商品化提供了理论支撑。A novel immunochromatic method for rapid detection of C.sakazakii was established in this paper.Antibody-modified magnetic beads(MNPs-IgG) were used as capture probes to separate and enrich C.sakazakii,which were then bound to biotin-labeled C.sakazakii antibody(Biotin-IgG) and avidin-peroxidase(Avidin-HRP).The resulting complex can react with the substrate 3,3’,5,5’-Tetramethylbenzidine(TMB).After the termination of the sulfuric acid,a linear relationship between the glucose concentration and the logarithm of the C.sakazakii concentration is obtained in the concentration range from 103-107 CFU/mL,and the regression equation is y =0.2353 x-0.2307.In this study,a new method based on colorimetric determination of C.sakazakii was successfully designed and optimized.The LOD of 8.68 ×10^2 CFU/mL is calculated according to the IUPAC definition(S/N=3) and a spiked sample detection rate of 96%.The detection time does not exceed 2.5 h,which provides theoretical support for practical application and commercialization.

关 键 词：信号转导探针 免疫磁性纳米颗粒 生物素亲和素系统 免疫比色法 食源性致病菌 

分 类 号：TS207.4[轻工技术与工程—食品科学]