【摘要】 基于阿卡斑病毒(Akabane virus, AKAV)S片段设计特异性引物和探针，并添加相应的基团将重组酶聚合酶等温扩增(recombinase polymerase amplification, RPA)与侧流层析试纸(lateral flow dipstick, LFD)相结合，通过对反应的温度、时间、引物及探针浓度进行优化，建立检测AKAV的RPA-LFD方法，并对其特异性、灵敏性和临床应用的可靠性进行评价。结果显示，该方法在37℃条件下反应20 min,即可在LFD试纸上读取检测结果，对蓝舌病病毒、牛多杀性巴氏杆菌、牛传染性鼻气管炎病毒和牛支原体均无交叉反应，最低可检出2.5×10~0拷贝/μL的标准质粒；临床样本检测结果与RT-PCR方法完全一致。结果表明，本试验所建立的RPA-LFD方法具有特异性好、灵敏度高、操作简便和可视化等优点，能够应用于临床检测，为AKAV的迅速确诊及防控提供新的技术支持。更多还原

【Abstract】 To establish a rapid visual detection method for Akabane virus(AKAV) on site, specific primers and probes based on the S fragment of AKAV were designed in this experiment.Corresponding groups were added to the primers or probes to fulfil the requirement of the combination of recombinase polymerase amplification(RPA) with lateral flow dipstick(LFD).The reaction temperature and time, concentrations of the primer and probe were optimized to establish the RPA-LFD method for detecting AKAV.After that, the specificity, sensitivity and clinical reliability of the method were evaluated.The results showed that after 20 minutes of reaction at 37 ℃,the test results could be read on LFD paper.There was no cross reaction against blue tongue virus, Pasteurella multocida,bovine infectious rhinotracheitis virus and bovine Mycoplasma bovis,and the detection limit was 2.5×10~0 copies/μL of standard plasmid.Detection of clinical samples showed a consistent results with that by RT-PCR method.These findings indicated that the RPA-LFD method established had the advantages of good specificity, high sensitivity, simple operation and visualization, and could be applied to clinical detection, which provides new technical support for the rapid diagnosis and prevention and control of AKAV.更多还原