基于一体化系统流感病毒快速检测技术的建立

Establishment of rapid influenza virus detection technology based on an integrated system

目的本研究旨在开发一种快速、准确的流感病毒一体化核酸检测方法。方法设计了针对近年来国内流感病毒主要流行株的特异性引物和探针,将其与提取、扩增试剂和即时检测(point of care test,POCT)系统整合,实现针对流感病毒的核酸提取、反应体系配制、扩增反应和结果判读功能的快速一体化病毒核酸检测流程。使用培养的流感病毒对检测体系进行检测下限测试,并对检测体系进行特异性测试。结果成功设计了甲型和乙型流感病毒的二重扩增引物和探针,实现了500拷贝/ml的检测下限,并通过特异性测试验证了检测试剂对腺病毒、呼吸道合胞病毒等其他呼吸道病原体无交叉反应。本研究建立的基于POCT技术的流感病毒核酸快速检测方法,可以实现在50 min内完成核酸提取、扩增和结果判读检测全流程,同时可以实现检测数据的实时上传。结论开发的基于POCT技术的流感病毒快速检测试剂盒,实现了"样品进、结果出"的便捷操作,适合在基层医疗机构中快速检测流感病毒,具有重要的临床应用价值。

Objective This study aimed to develop a rapid and accurate integrated nucleic acid detection method tailored for the influenza virus.Methods We designed primers and probes targeting the predominant influenza virus strains circulating in China in recent years.These were integrated with extraction and amplification reagents and a point of care testing(POCT)system to facilitate a seamless and expedited process involving nucleic acid extraction,reaction system preparation,amplification,and result interpretation for the influenza virus.The specificity of the POCT system was evaluated using cultured influenza viruses,while its cross-reactivity was assessed against common respiratory pathogens,including adenovirus and respiratory syncytial virus.Results Our study successfully developed duplex amplification primers and probes for both influenza A and B viruses,achieving a detection threshold as low as 500 copies/ml.Specificity tests confirmed that the detection reagents did not show cross-reactivity with other respiratory pathogens such as adenovirus and respiratory syncytial virus.The POCT-based rapid nucleic acid detection method for influenza virus was established,it is capable of completing the entire process from nucleic acid extraction to amplification and result interpretation within 50 minutes,while enabling real-time data upload.Conclusions The POCT-based rapid influenza virus detection kit developed in this study offers a"sample in,results out"convenience,making it suitable for rapid influenza virus detection in primary care settings.This innovation has significant potential for clinical application.