基于核酸等温扩增的病原微生物微流控检测技术

Methods of isothermal nucleic acid amplification-based microfluidic chips for pathogen microorganism detection

病原微生物的快速检测对疫情的预防控制至关重要。基于PCR的病原微生物核酸检测方法克服了传统病原微生物培养方法耗时长、免疫学检测存在窗口期等问题,已成为目前最主要的病原微生物筛查方法。然而,对精确控温热循环仪的依赖却严重限制了其在资源匮乏地区的应用。虽然基于核酸等温扩增的病原微生物检测方法可摆脱对高精度温控设备的依赖,但仍需要进行样本核酸分离提取、扩增与检测等步骤。近年来,微流控技术与核酸等温扩增技术相结合,诞生了多种病原微生物等温扩增微流控检测技术。该技术通过设计芯片结构、优化进样模式及检测方式,实现了病原微生物核酸提取、扩增与检测一体化,并具备多重检测、定量检测等功能,具有对仪器依赖度小、对操作人员要求不高、样本量需求小和自动化程度高等优点,适合于在多种环境下的病原微生物快速检测。本文从核酸等温扩增原理、进样方式、检测方式等方面对核酸等温扩增病原微生物微流控检测技术进行了综述,以期为病原微生物的快速筛查提供更多的方案思路,提升公共卫生领域对传染性疾病的防控能力。

Rapid detection of pathogenic microorganisms is key to the epidemiologic identification,prevention and control of disease in the field of public health.PCR-based pathogen detection methods have been widely used because they overcome the time-consuming issues that traditional culture-based methods required including the limited window required by immunological detection.However,the requirement on precision temperature-controlled thermal cyclers severely limits their use in resource-limited areas.The detection methods of pathogenic microorganisms based on isothermal amplification of nucleic acids are free of dependence on high-precision temperature control equipment,but requirements for nucleic acids extraction,amplification and detection must be defined.In recent years,a number of alternative methods for pathogenic microorganism detection have been developed by combining microfluidic technology with nucleic acid isothermal amplification technology.By designing the chip structures,optimizing the injection modes,and utilizing multiple detection and quantitative methods,the integration of pathogen nucleic acid extraction,amplification and detection is achieved.The method provides advantages of less instrument dependence,decreased operator requirements,smaller sample size,and higher automation which are suitable for the rapid detection of pathogenic microorganisms in various environments.In this review,we summarize several microfluidic detection methods based on nucleic acid isothermal amplification for pathogens including amplification principles,injection methods and detection methods.These methods provide more capability for the rapid screening of pathogenic microorganisms which enhances the management of infectious diseases in the field of public health.