食品科学 ›› 2018, Vol. 39 ›› Issue (6): 302-306.doi: 10.7506/spkx1002-6630-201806047

• 安全检测 • 上一篇    下一篇

流式分析技术快速定量检测牛乳中大肠杆菌O157:H7

刘思渊1,2,古少鹏1,*,隋志伟2,*,刘思章2,薛蕾2,霍乃蕊1   

  1. (1.山西农业大学动物科技学院,山西?太谷 030801;2.中国计量科学研究院,北京 100013)
  • 出版日期:2018-03-25 发布日期:2018-03-14
  • 基金资助:
    山西省重点研发计划重点项目(201603D21108-03);“十三五”国家重点研发计划重点专项(2017YFF0204602);国家标准物质资源共享平台项目(32-APT1702-9)

Rapid and Quantitative Detection of Escherichia coli O157:H7 in Milk by Flow Cytometry

LIU Siyuan1,2, GU Shaopeng1,*, SUI Zhiwei2,*, LIU Sizhang2, XUE Lei2, HUO Nairui1   

  1. (1. College of Animal Science and Veterinary Medicine, Shanxi Agricultural University, Taigu 030801, China; 2. National Institute of Metrology, Beijing 100013, China)
  • Online:2018-03-25 Published:2018-03-14

摘要: 建立一种基于流式分析技术的快速定量检测牛乳中大肠杆菌O157:H7的方法。用偶联有异硫氰酸荧光素的大肠杆菌O157单克隆抗体对大肠杆菌O157:H7进行特异性标记,通过优化抗体反应条件,建立流式检测方法,然后对磷酸盐缓冲溶液(phosphate buffer saline,PBS)和人工污染牛乳样品中不同浓度的大肠杆菌O157:H7进行定量检测。本研究建立的流式检测方法的在PBS中的检测范围为2.57×103~1.12×108?CFU/mL,灵敏度达到2.57×103?CFU/mL。将所建立的流式检测方法应用于牛乳样品检测,当人工污染牛乳样品中大肠杆菌O157:H7的浓度在2.31×104~1.48×108?CFU/mL之间时,流式检测方法与平板计数方法检测结果基本一致,方法的灵敏度为2.31×104?CFU/mL,检测时间为35?min。该方法能快速、定量地检测出牛乳样品中的大肠杆菌O157:H7,在食源性致病菌的快速筛查和监控中具有重要的应用价值。

关键词: 流式分析技术, 快速检测, 大肠杆菌O157:H7, 牛乳

Abstract: The purpose of this study was to develop a flow cytometry (FCM) method for rapid and quantitative detection of Escherichia coli O157:H7 in milk. E. coli O157:H7 was labeled with fluorescein isothiocyanate (FITC)-conjugated anti-E. coli O157 monocolonal antibodies and then detected by flow cytometry (green fluorescence channel). Serial 10-fold dilutions of E. coli O157:H7 in PBS were tested by FCM method under optimized reaction conditions. The detection range of this method was 2.57 × 103–1.12 × 108 CFU/mL and the sensitivity was 2.57 × 103 CFU/mL. Then the FCM method was applied to artificially contaminated milk samples with 2.31 × 104–1.48 × 108 CFU/mL E. coli O157:H7. The results were consistent with the plate counting method. The sensitivity of our method was 2.31 × 104 CFU/mL and the process required 35 min. This method allowed for rapid, efficient and sensitive detection of E. coli O157:H7 in milk samples. Thus, it has a great potential for application in rapid screening and monitoring of foodborne pathogens.

Key words: flow cytometry, rapid detection, Escherichia coli O157:H7, milk

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