Shelby Tyler, Sulthana Shoukath, McAfee James, Banerjee Tuhina, Santra Santimukul
Department of Chemistry and Kansas Polymer Research Center, Pittsburg State University.
Department of Chemistry and Kansas Polymer Research Center, Pittsburg State University;
J Vis Exp. 2017 Sep 17(127):55821. doi: 10.3791/55821.
Enterohemorrhagic Escherichia coli O157:H7 has been linked to both waterborne and foodborne illnesses, and remains a threat despite the food- and water-screening methods used currently. While conventional bacterial detection methods, such as polymerase chain reaction (PCR) and enzyme-linked immunosorbent assays (ELISA) can specifically detect pathogenic contaminants, they require extensive sample preparation and lengthy waiting periods. In addition, these practices demand sophisticated laboratory instruments and settings, and must be executed by trained professionals. Herein, a protocol is proposed for a simpler diagnostic technique that features the unique combination of magnetic and fluorescent parameters in a nanoparticle-based platform. The proposed multiparametric magneto-fluorescent nanosensors (MFnS) can detect E. coli O157:H7 contamination with as little as 1 colony-forming unit present in solution within less than 1 h. Furthermore, the ability of MFnS to remain highly functional in complex media such as milk and lake water has been verified. Additional specificity assays were also used to demonstrate the ability of MFnS to only detect the specific target bacteria, even in the presence of similar bacterial species. The pairing of magnetic and fluorescent modalities allows for the detection and quantification of pathogen contamination in a wide range of concentrations, exhibiting its high performance in both early- and late-stage contamination detection. The effectiveness, affordability, and portability of the MFnS make them an ideal candidate for point-of-care screening for bacterial contaminants in a wide range of settings, from aquatic reservoirs to commercially packaged foods.
肠出血性大肠杆菌O157:H7与水源性和食源性疾病都有关联,尽管目前采用了食品和水筛查方法,但它仍然构成威胁。虽然传统的细菌检测方法,如聚合酶链反应(PCR)和酶联免疫吸附测定(ELISA)能够特异性地检测致病污染物,但它们需要大量的样品制备和漫长的等待时间。此外,这些操作需要精密的实验室仪器和环境,并且必须由经过培训的专业人员执行。在此,我们提出了一种更简单的诊断技术方案,该方案在基于纳米颗粒的平台中结合了磁性和荧光参数的独特组合。所提出的多参数磁荧光纳米传感器(MFnS)能够在不到1小时内检测出溶液中低至1个菌落形成单位的大肠杆菌O157:H7污染。此外,已经验证了MFnS在牛奶和湖水等复杂介质中仍能保持高功能。还使用了额外的特异性检测来证明MFnS即使在存在相似细菌种类的情况下也只能检测特定目标细菌的能力。磁性和荧光模式的结合使得能够检测和定量各种浓度的病原体污染,在早期和晚期污染检测中均表现出高性能。MFnS的有效性、可负担性和便携性使其成为在从水库到商业包装食品等广泛环境中进行细菌污染物即时检测筛查的理想候选者。