Department of Microbiology, Immunology, and Infectious Diseases, University of Calgary, Calgary, AB, Canada.
Department of Mechanical and Manufacturing Engineering, University of Calgary, Calgary, AB, Canada.
Sci Rep. 2021 Jun 28;11(1):13378. doi: 10.1038/s41598-021-92677-z.
The highly infectious nature of SARS-CoV-2 necessitates the use of widespread testing to control the spread of the virus. Presently, the standard molecular testing method (reverse transcriptase-polymerase chain reaction, RT-PCR) is restricted to the laboratory, time-consuming, and costly. This increases the turnaround time for getting test results. This study sought to develop a rapid, near-patient saliva-based test for COVID-19 (Saliva-Dry LAMP) with similar accuracy to that of standard RT-PCR tests. A lyophilized dual-target reverse transcription-loop-mediated isothermal amplification (RT-LAMP) test with fluorometric detection by the naked eye was developed. The assay relies on dry reagents that are room temperature stable. A device containing a centrifuge, heat block, and blue LED light system was manufactured to reduce the cost of performing the assay. This test has a limit of detection of 1 copy/µL and achieved a positive percent agreement of 100% [95% CI 88.43% to 100.0%] and a negative percent agreement of 96.7% [95% CI 82.78-99.92%] relative to a reference standard test. Saliva-Dry LAMP can be completed in 105 min. Precision, cross-reactivity, and interfering substances analysis met international regulatory standards. The combination of ease of sample collection, dry reagents, visual detection, low capital equipment cost, and excellent analytical sensitivity make Saliva-Dry LAMP particularly useful for resource-limited settings.
SARS-CoV-2 具有高度传染性，因此需要广泛检测以控制病毒传播。目前，标准的分子检测方法（逆转录-聚合酶链反应，RT-PCR）受到实验室条件、耗时和昂贵的限制。这增加了获得检测结果的周转时间。本研究旨在开发一种快速、接近患者的基于唾液的 COVID-19 检测方法（Saliva-Dry LAMP），其准确性与标准 RT-PCR 检测相当。开发了一种冻干双靶标逆转录环介导等温扩增（RT-LAMP）检测，通过肉眼进行荧光检测。该检测依赖于室温稳定的干试剂。制造了一种包含离心机、加热块和蓝色 LED 灯系统的设备，以降低执行检测的成本。该检测的检测限为 1 拷贝/µL，与参考标准检测相比，阳性符合率为 100%[95%置信区间 88.43%至 100.0%]，阴性符合率为 96.7%[95%置信区间 82.78%至 99.92%]。Saliva-Dry LAMP 可在 105 分钟内完成。精密度、交叉反应性和干扰物质分析符合国际监管标准。样本采集简便、干试剂、目视检测、低资本设备成本和出色的分析灵敏度相结合，使 Saliva-Dry LAMP 特别适用于资源有限的环境。
