Department of Clinical Laboratory, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, Guangzhou, 510120, China.
Shenzhen Key Laboratory of Ophthalmology, Ocular Trauma Treatment and Stem Cell Differentiation Public Service Platform of Shenzhen, Shenzhen Eye Hospital, Shenzhen, 518040, China.
Bioprocess Biosyst Eng. 2021 May;44(5):977-984. doi: 10.1007/s00449-020-02423-4. Epub 2020 Aug 29.
During a large variety of common pathogens, E. coli, P. aeruginosa, MRSA, MRCNS, V. parahaemolyticus, L. monocytogenes and Salmonella are the leading pathogens responsible for large number of human infections and diseases. In this study, a high flux screening based on nucleic acid isothermal amplification technique has been developed. For the 8 common pathogens, species-specific targets had been selected and analyzed for their unique specificity. After optimization, separate LAMP reaction assays had been bioprocessed and integrated into one systematic detection platform, including 8 strips (PCR tubes) and 96-well plates. Eight standard strains verified for the accuracy. Application of the established high flux screening platform was used for detection for 48 samples in 4 different 96-well plates, with 2 groups of 2 operators using double-blind procedure. The accuracy of 100% was obtained, with the total time consumption as 66-75 min (for 12 samples detection on 8 different pathogens). As concluded, through the bioprocess of the systematic platform based on LAMP technique, it's been demonstrated to be capable of simultaneous detection of 8 pathogens, with high sensitivity, specificity, rapidity and convenience.
在多种常见病原体中，大肠杆菌、铜绿假单胞菌、耐甲氧西林金黄色葡萄球菌、耐甲氧西林凝固酶阴性葡萄球菌、副溶血性弧菌、单核细胞增生李斯特菌和沙门氏菌是导致大量人类感染和疾病的主要病原体。在本研究中，开发了一种基于核酸等温扩增技术的高通量筛选方法。针对这 8 种常见病原体，选择了物种特异性靶标，并对其独特的特异性进行了分析。经过优化后，单独的 LAMP 反应分析已被生物加工并集成到一个系统检测平台中，包括 8 个条（PCR 管）和 96 孔板。8 个标准菌株用于验证准确性。建立的高通量筛选平台用于检测 4 个不同 96 孔板中的 48 个样本，由 2 组操作员使用双盲程序进行。获得了 100%的准确率，总耗时为 66-75 分钟（用于检测 8 种不同病原体的 12 个样本）。综上所述，通过基于 LAMP 技术的系统平台的生物加工，证明其能够同时检测 8 种病原体，具有高灵敏度、特异性、快速性和便利性。
