Master Program of Biotechnology, Institute for Research and Medical Consultations, Imam Abdulrahman Bin Faisal University, Dammam, 31441, Saudi Arabia.
Department of Genetics Research, Institute for Research and Medical Consultations (IRMC), Imam Abdulrahman Bin Faisal University, Dammam, 31441, Saudi Arabia.
Sci Rep. 2023 Mar 28;13(1):5066. doi: 10.1038/s41598-023-31760-z.
The reverse-transcription loop-mediated isothermal amplification (RT-LAMP) is a cheaper and faster testing alternative for detecting SARS-CoV-2. However, a high false-positive rate due to misamplification is one of the major limitations. To overcome misamplifications, we developed colorimetric and fluorometric RT-LAMP assays using five LAMP primers, instead of six. The gold-standard RT-PCR technique verified the assays' performance. Compared to other primer sets with six primers (N, S, and RdRp), the E-ID1 primer set, including five primers, performed superbly on both colorimetric and fluorometric assays. The sensitivity of colorimetric and fluorometric assays was 89.5% and 92.2%, respectively, with a limit of detection of 20 copies/µL. The colorimetric RT-LAMP had a specificity of 97.2% and an accuracy of 94.5%, while the fluorometric RT-LAMP obtained 99% and 96.7%, respectively. No misamplification was evident even after 120 min, which is crucial for the success of this technique. These findings are important to support the use of RT-LAMP in the healthcare systems in fighting COVID-19.
逆转录环介导等温扩增（RT-LAMP）是一种更廉价、更快速的 SARS-CoV-2 检测替代方法。然而，由于错误扩增导致的高假阳性率是其主要限制之一。为了克服错误扩增，我们使用五种 LAMP 引物而不是六种开发了比色和荧光 RT-LAMP 检测方法。金标准 RT-PCR 技术验证了这些检测方法的性能。与其他使用六种引物（N、S 和 RdRp）的引物组相比，包括五种引物的 E-ID1 引物组在比色和荧光检测中表现出色。比色和荧光检测的灵敏度分别为 89.5%和 92.2%，检测限为 20 拷贝/µL。比色 RT-LAMP 的特异性为 97.2%，准确性为 94.5%，而荧光 RT-LAMP 的特异性和准确性分别为 99%和 96.7%。即使在 120 分钟后也没有明显的错误扩增，这对该技术的成功至关重要。这些发现对于支持 RT-LAMP 在抗击 COVID-19 的医疗保健系统中的应用非常重要。
