Magnetic Nanoparticle-Based Platform for Characterization of Shiga-like Toxin 1 from Complex Samples

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• 作者：Fang-Yin Kuo ; Bo-Yao Chang ; Ching-Yi Wu ; Kwok-Kong Tony Mong ; Yu-Chie Chen
• 刊名：Analytical Chemistry
• 出版年：2015
• 出版时间：October 20, 2015
• 年：2015
• 卷：87
• 期：20
• 页码：10513-10520
• 全文大小：395K
• ISSN：1520-6882

文摘

Foodborne illness outbreaks resulting from contamination of Escherichia coli O157:H7 remain a serious concern in food safety. E. coli O157:H7 can cause bloody diarrhea, hemolytic uremic syndrome, or even death. The pathogenicity of E. coli O157:H7 is mainly caused by the expression of Shiga-like toxins (SLTs), i.e., SLT-1 and SLT-2. SLTs are pentamers composed of a single A and five B subunits. In this study, we propose a magnetic nanoparticle (MNP)-based platform to rapidly identify SLT-1 from the complex cell lysate of E. coli O157:H7. The core of the MNPs is made of iron oxide, whereas the surface of the core is coated with a thin layer of alumina (Fe₃O₄@Al₂O₃ MNPs). The Fe₃O₄@Al₂O₃ MNPs are functionalized with pigeon ovalbumin (POA), which contains Gal-伪(1鈫?)-Gal-尾(1鈫?)-GlcNAc termini that can bind SLT-1B selectively. Furthermore, POA is a phosphate protein. Thus, it can be easily immobilized on the surface of the Fe₃O₄@Al₂O₃ MNPs through aluminum phosphate chelation under microwave heating within 1.5 min. The generated POA鈥揊e₃O₄@Al₂O₃ MNPs are capable of effectively enriching SLT-1B from complex cell lysates simply by pipetting 20 渭L of the sample in and out of the tip in a vial for 鈭? min. To release SLT-1 from the MNPs, Gal-伪(1鈫?)-Gal disaccharides were used for displacement. The released target species are sufficient to be identified by matrix-assisted laser desorption/ionization mass spectrometry. Although the sample volume used in this approach is small (20 渭L) and the enrichment time is short (1 min), the selectivity of this approach toward SLT-1B is quite good. We have demonstrated the effectiveness of this approach for rapid determination of the presence of SLT-1 from complex cell lysates and ham/juice samples based on the detection of SLT-1B.