食品中金黄色葡萄球菌PCR-ELISA检测技术建立  

Establishment of PCR-ELISA technology for Staphylococcus aureus in food

作　　者：胡金强[1,2,3] 雷俊婷 白艳红[1] 魏向珂 景建洲[1] 高辉[1] 孙新城[1] 董彩文[1] 耿尧[1] 姜春鹏[1] 

机构地区：[1]郑州轻工业学院食品与生物工程学院,河南郑州450002 [2]河南省食品安全国际联合实验室,河南郑州450002 [3]食品生产与安全河南省协同创新中心,河南郑州450002

出　　处：《食品工业科技》2016年第20期63-67,共5页Science and Technology of Food Industry

基　　金：国家自然科学基金(31201901);教育部留学回国人员科研启动基金(第46批;教外司留[2013]693号);河南省高校青年骨干教师资助计划项目(教高[2015]1032号);河南省教育厅科学技术重点研究项目(14A180025);郑州市科技攻关项目(20130857);郑州轻工业学院博士基金项目(2011BSJJ033);郑州轻工业学院青年骨干教师资助计划项目(2013QNGG02);郑州轻工业学院研究生创新基金项目(2014031);食品生产与安全河南省协同创新中心研究生创新基金项目(FCICY201614)

摘　　要：针对食品中金黄色葡萄球菌检测,选取金黄色葡萄球菌耐热核酸酶nuc基因作为靶标,利用地高辛标记引物扩增的PCR产物与生物素标记的探针杂交,然后与ELISA平板上的链霉素杂交,通过抗地高辛抗体显色建立PCR-ELISA检测技术。应用该方法检测人工污染牛奶中金黄色葡萄球菌,同时将大肠埃希氏菌、沙门氏菌、志贺氏菌、单增李斯特菌等常见食源性致病菌作为阴性对照。结果显示,金黄色葡萄球菌的纯培养物以及人工污染金黄色葡萄球菌牛奶,金黄色葡萄球菌为阳性,其他对照菌为阴性,两者PCR-ELISA检测敏感性均为10-1CFU/m L,比普通PCR检测的敏感性（10-3CFU/m L）高100倍。因此,该研究建立的PCR-ELISA方法具有良好的特异性与敏感性,能够特异、敏感、准确地检测牛奶中的金黄色葡萄球菌,为食品中金黄色葡萄菌的快速鉴定、风险评估与有效监测提供重要技术手段。The nuc gene was as a target to detect Staphylococcus aureus in food. The PCR products were hybridized with digoxin-labeled probes, and then with streptomycin in microwell plate. PCR-ELISA method had been established by chromogenic reaction with anti- digoxin antibody. This method was applied to detect Staphylococcus aureus in milk artificially contaminated with Staphylococcus aureus,while other bacteria strains, including Escherichia coil, Salmonella, Shigella, Listeria monocytogenes were as negative controls. The results showed that for the pure culture and artificially contaminated milk, Staphylococcus aureus were positive, while negative for negative controls. The sensitivity of detection of Staphylococcus aureus for the pure culture and artificially contaminated milk were 10^1 CFU/mL, a 100- fold higher than that of conventional PCR method （103 CFU/mL）. Therefore,PCR-ELISA established in this study for detection of Staphylococcus aureus in milk sample was specifically, sensitively and accurately, which should provide an important technical means for rapid identification,risk assessment and effective monitoring of Staphylococcus aureus in food.

关 键 词：金黄色葡萄球菌 PCR-ELISA 检测 

分 类 号：TS207.3[轻工技术与工程—食品科学]