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开发一种用于同时简单检测食源性病原体的磁性纳米颗粒微阵列。

Development of a magnetic nanoparticles microarray for simultaneous and simple detection of foodborne pathogens.

机构信息

Hunan Key Laboratory of Green Packaging and Application of Biological Nanotechnology, Hunan University of Technology, Zhuzhou 412007, PR China.

出版信息

J Biomed Nanotechnol. 2013 Jul;9(7):1254-60. doi: 10.1166/jbn.2013.1610.

Abstract

Foodborne diseases are a widespread and growing public health problem affecting both developed and developing countries, microbiologically contaminated food and water are the major causes of diarrhoeal diseases. Methods based on polymerase chain reaction (PCR) and microarrays are rapid and sensitive enough to detect very small quantities of microorganisms, however, the requirement for expensive equipments limits their application. In the present paper, we describe a method based on multiplex PCR and magnetic nanoparticles labelling for simultaneous detection of four major foodborne pathogens, including Escherichia coli O157:H7, Salmonella enterica, Vibrio cholera and Campylobacter jejuni. The process utilizes an oligonucleotide array onto which 5' biotinylated single strand PCR products were hybridized and visualized with streptavidin-coated magnetic nanoparticles (SA-MNPs), the signal from which could be detected by the naked eye, microscope or CCD camera. By employing SA-MNPs as visible labels, the microarray unambiguously distinguished all 4 pathogens with detection sensitivity up to 316 CFU/mL. Due to its high sensitivity, specificity and simple detection procedure, the magnetic bead assay provides a powerful tool for the detection and identification of foodborne pathogens in a modestly equipped laboratory.

摘要

食源性疾病是一个广泛且日益严重的公共卫生问题,影响着发达国家和发展中国家。受微生物污染的食物和水是导致腹泻病的主要原因。基于聚合酶链反应(PCR)和微阵列的方法足够快速和灵敏,可检测到非常少量的微生物,但昂贵的设备要求限制了它们的应用。在本文中,我们描述了一种基于多重 PCR 和磁性纳米粒子标记的方法,用于同时检测四种主要的食源性致病菌,包括大肠杆菌 O157:H7、沙门氏菌、霍乱弧菌和空肠弯曲菌。该过程利用寡核苷酸阵列,将 5'生物素化单链 PCR 产物杂交,并与链霉亲和素包被的磁性纳米粒子(SA-MNPs)可视化,其信号可以通过肉眼、显微镜或 CCD 相机检测。通过将 SA-MNPs 用作可见标记,微阵列可以明确地区分所有 4 种病原体,检测灵敏度高达 316 CFU/mL。由于其高灵敏度、特异性和简单的检测程序,磁性珠测定法为在设备简陋的实验室中检测和鉴定食源性致病菌提供了有力工具。

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