Wang Meng, Chen Jianjun, Zhang Zhi-Ling
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan, Hubei 430072, China.
Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Center for Biosafety Mega-Science, Chinese Academy of Sciences, Wuhan, Hubei 430071, China.
Anal Chem. 2024 Sep 11. doi: 10.1021/acs.analchem.4c03052.
Influenza A virus (IAV) can cause infectious respiratory diseases in humans and animals. IAVs mutate rapidly through antigenic drift and shift, resulting in the emergence of numerous IAV subtypes and significant challenges for IAV detection. Therefore, achieving the simultaneous detection of multiple IAVs is crucial. In this work, three specific aptamers targeting the hemagglutination (HA) protein of the influenza A H5N1, H7N9, and H9N2 viruses were screened using a multichannel magnetic microfluidic chip. The aptamers exhibit nanomolar affinity and excellent specificity for the HA protein of H5N1, H7N9, and H9N2 viruses. Furthermore, three specific aptamers were truncated and labeled with different fluorescence markers to realize fluorescence quantitative detection of influenza A H5N1, H7N9, and H9N2 viruses through an aptamer sandwich assay in 1 h. The limit of detection (LOD) of the developed method is 0.38 TCID/mL for the H5N1 virus, 0.75 TCID/mL for the H7N9 virus, and 1.14 TCID/mL for the H9N2 virus. The detection method has excellent specificity, strong anti-interference ability, and good reproducibility. This work provides a sensitive quantitative detection method for the H5N1, H7N9, and H9N2 viruses, enabling quantitative fluorescence detection for multiple IAV subtypes.
甲型流感病毒（IAV）可导致人类和动物的传染性呼吸道疾病。IAV通过抗原漂移和转换迅速变异，导致众多IAV亚型出现，并给IAV检测带来重大挑战。因此，实现多种IAV的同时检测至关重要。在这项工作中，使用多通道磁性微流控芯片筛选了三种靶向甲型H5N1、H7N9和H9N2流感病毒血凝素（HA）蛋白的特异性适体。这些适体对H5N1、H7N9和H9N2病毒的HA蛋白表现出纳摩尔亲和力和优异的特异性。此外，对三种特异性适体进行截短并标记不同的荧光标记物，以通过适体夹心测定在1小时内实现对甲型H5N1、H7N9和H9N2流感病毒的荧光定量检测。所开发方法对H5N1病毒的检测限（LOD）为0.38 TCID/mL，对H7N9病毒为0.75 TCID/mL，对H9N2病毒为1.14 TCID/mL。该检测方法具有优异的特异性、较强的抗干扰能力和良好的重现性。这项工作为H5N1、H7N9和H9N2病毒提供了一种灵敏的定量检测方法，能够对多种IAV亚型进行定量荧光检测。
