环介导等温扩增
肉眼
铜
检出限
固相萃取
DNA
材料科学
屎肠球菌
DNA提取
化学
色谱法
聚合酶链反应
冶金
生物化学
抗生素
基因
作者
Hanh An Nguyen,Nae Yoon Lee
标识
DOI:10.1016/j.bios.2023.115222
摘要
In this study, an all-in-one poly(methyl methacrylate) (PMMA) device integrating two novel techniques – DNA extraction employing a CuSO4/H2O2 system and DNA detection utilizing solid phase copper tape – coupled with loop-mediated isothermal amplification (LAMP) is developed for on-site pathogen detection. The CuSO4/H2O2 system, also known as Fenton-like reaction, is used to produce hydroxyl radicals, which can disrupt bacterial membranes via lipid peroxidation and release DNA at room temperature. The released DNA is subsequently amplified by LAMP reaction. The acidic environment resulting from the production of hydrogen ions in the presence of target DNA in the LAMP reaction can stimulate the color change on copper tape due to the corrosion, while the innate alkaline environment in a negative sample not containing target DNA cannot stimulate the corrosion. The fabricated PMMA device integrates all the functionalities necessary for molecular diagnostics such as DNA extraction, amplification, and detection, and a carbon paste-based heater is fabricated for LAMP reaction. Using the PMMA device, Enterococcus faecium was detected as low as 4.67 × 102 CFU/mL within 90 min. E. faecium spiked in milk was successfully detected using the all-in-one PMMA device. The equipment-free techniques for decentralized diagnostics and naked-eye readout of results coupled with the portable heater serves as a promising solution for point-of-care testing particularly in a resource-limited environment.
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