用于检测唾液中甲型H1N1流感病毒的集成微流控预浓缩和核酸扩增系统

Integrated Microfluidic Preconcentration and Nucleic Amplification System for Detection of Influenza A Virus H1N1 in Saliva.

作者信息

Kim Yonghee, Abafogi Abdurhaman Teyib, Tran Buu Minh, Kim Jaewon, Lee Jinyeop, Chen Zhenzhong, Bae Pan Kee, Park Kyoungsook, Shin Yong-Beom, van Noort Danny, Lee Nae Yoon, Park Sungsu

机构信息

School of Mechanical Engineering, Sungkyunkwan University, Suwon 16419, Korea.

Department of BioNano Technology, College of BioNano Technology, Gachon University, Seongnam 13120, Korea.

出版信息

Micromachines (Basel). 2020 Feb 16;11(2):203. doi: 10.3390/mi11020203.

Influenza A viruses are often present in environmental and clinical samples at concentrations below the limit of detection (LOD) of molecular diagnostics. Here we report an integrated microfluidic preconcentration and nucleic amplification system (μFPNAS) which enables both preconcentration of influenza A virus H1N1 (H1N1) and amplification of its viral RNA, thereby lowering LOD for H1N1. H1N1 virus particles were first magnetically preconcentrated using magnetic nanoparticles conjugated with an antibody specific for the virus. Their isolated RNA was amplified to cDNA through thermocycling in a trapezoidal chamber of the μFPNAS. A detection limit as low as 100 TCID50 (50% tissue culture infective dose) in saliva can be obtained within 2 hours. These results suggest that the LOD of molecular diagnostics for virus can be lowered by systematically combining immunomagnetic separation and reverse transcriptase-polymerase chain reaction (RT-PCR) in one microfluidic device.

甲型流感病毒在环境和临床样本中的浓度往往低于分子诊断的检测限（LOD）。在此，我们报告了一种集成微流控预浓缩和核酸扩增系统（μFPNAS），该系统能够对甲型H1N1流感病毒（H1N1）进行预浓缩并扩增其病毒RNA，从而降低H1N1的检测限。首先，使用与该病毒特异性抗体偶联的磁性纳米颗粒对H1N1病毒颗粒进行磁性预浓缩。在μFPNAS的梯形腔室中通过热循环将其分离的RNA扩增为cDNA。在2小时内可在唾液中获得低至100个TCID50（50%组织培养感染剂量）的检测限。这些结果表明，通过在一个微流控装置中系统地结合免疫磁分离和逆转录聚合酶链反应（RT-PCR），可以降低病毒分子诊断的检测限。