Fry Jacob, Lee Jean Y H, McAuley Julie L, Porter Jessica L, Monk Ian R, Martin Samuel T, Collins David J, Barbante Gregory J, Fitzgerald Nicholas J, Stinear Timothy P
ARC Centre of Excellence in Exciton Science, The School of Chemistry, The University of Melbourne, Masson Rd, Parkville, Victoria 3010, Australia.
Department of Microbiology and Immunology, The Doherty Institute for Infection and Immunity, The University of Melbourne, 792 Elizabeth Street, Melbourne, Victoria 3000, Australia.
ACS Omega. 2024 Sep 20;9(39):40832-40840. doi: 10.1021/acsomega.4c05784. eCollection 2024 Oct 1.
The Coronavirus disease 2019 (COVID-19) pandemic has supercharged innovation in the field of molecular diagnostics and led to the exploration of systems that permit the autonomous identification of airborne infectious agents. Airborne virus detection is an emerging approach for determining exposure risk, although current methods limit intervention timeliness. Here, we explore reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays for one-pot detection of Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) (SCV2) run on membrane filters suitable for micro-air-filtration of airborne viruses. We use a design of experiments statistical framework to establish the optimal additive composition for running RT-LAMP on membrane filters. Using SCV2 liquid spike-in experiments and fluorescence detection, we show that single-pot RT-LAMP on glass fiber filters reliably detected 0.10 50% tissue culture infectious dose (TCID) SCV2 per reaction (3600 E-gene copies) and is an order of magnitude more sensitive than conventional RT-LAMP.
2019年冠状病毒病（COVID-19）大流行极大地推动了分子诊断领域的创新，并促使人们探索能够自主识别空气传播感染源的系统。空气传播病毒检测是一种确定暴露风险的新兴方法，尽管目前的方法限制了干预的及时性。在这里，我们探索了逆转录环介导等温扩增（RT-LAMP）检测法，用于在适用于空气传播病毒微空气过滤的膜过滤器上进行一锅法检测严重急性呼吸综合征冠状病毒2（SARS-CoV-2）（SCV2）。我们使用实验设计统计框架来确定在膜过滤器上进行RT-LAMP的最佳添加剂组成。通过SCV2液体加标实验和荧光检测，我们表明在玻璃纤维过滤器上进行的单锅RT-LAMP能够可靠地检测到每个反应0.10 50%组织培养感染剂量（TCID）的SCV2（3600个E基因拷贝），并且比传统RT-LAMP敏感一个数量级。
