PT - JOURNAL ARTICLE AU - Dao Thi, Viet Loan AU - Herbst, Konrad AU - Boerner, Kathleen AU - Meurer, Matthias AU - Kremer, Lukas PM AU - Kirrmaier, Daniel AU - Freistaedter, Andrew AU - Papagiannidis, Dimitrios AU - Galmozzi, Carla AU - Stanifer, Megan L. AU - Boulant, Steeve AU - Klein, Steffen AU - Chlanda, Petr AU - Khalid, Dina AU - Barreto Miranda, Isabel AU - Schnitzler, Paul AU - Kräusslich, Hans-Georg AU - Knop, Michael AU - Anders, Simon TI - A colorimetric RT-LAMP assay and LAMP-sequencing for detecting SARS-CoV-2 RNA in clinical samples AID - 10.1126/scitranslmed.abc7075 DP - 2020 Aug 12 TA - Science Translational Medicine PG - eabc7075 VI - 12 IP - 556 4099 - http://stm.sciencemag.org/content/12/556/eabc7075.short 4100 - http://stm.sciencemag.org/content/12/556/eabc7075.full AB - We need simple methods to rapidly test large numbers of people for infection with the SARS-CoV-2 coronavirus. Quantitative PCR (qPCR) after reverse transcription (RT), the standard method, is very sensitive but requires expensive instrumentation. Loop-mediated isothermal amplification (LAMP) is an alternative to qPCR that is faster and requires fewer resources. Dao Thi et al. tested the RT-LAMP assay on several hundred clinical RNA samples isolated from pharyngeal swabs collected from individuals being tested for COVID-19. They confirmed that the RT-LAMP assay was a simpler albeit less sensitive option compared to RT-qPCR for large-scale testing for SARS-CoV-2 RNA. These investigators also developed a simplified version of this method (direct swab–to–RT-LAMP assay) that did not require a prior RNA isolation step as well as a method for highly multiplexed sequencing of RT-LAMP reactions (LAMP-sequencing).The coronavirus disease 2019 (COVID-19) pandemic caused by the SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) coronavirus is a major public health challenge. Rapid tests for detecting existing SARS-CoV-2 infections and assessing virus spread are critical. Approaches to detect viral RNA based on reverse transcription loop-mediated isothermal amplification (RT-LAMP) have potential as simple, scalable, and broadly applicable testing methods. Compared to RT quantitative polymerase chain reaction (RT-qPCR)–based methods, RT-LAMP assays require incubation at a constant temperature, thus eliminating the need for sophisticated instrumentation. Here, we tested a two-color RT-LAMP assay protocol for detecting SARS-CoV-2 viral RNA using a primer set specific for the N gene. We tested our RT-LAMP assay on surplus RNA samples isolated from 768 pharyngeal swab specimens collected from individuals being tested for COVID-19. We determined the sensitivity and specificity of the RT-LAMP assay for detecting SARS-CoV-2 viral RNA. Compared to an RT-qPCR assay using a sensitive primer set, we found that the RT-LAMP assay reliably detected SARS-CoV-2 RNA with an RT-qPCR cycle threshold (CT) number of up to 30, with a sensitivity of 97.5% and a specificity of 99.7%. We also developed a swab–to–RT-LAMP assay that did not require a prior RNA isolation step, which retained excellent specificity (99.5%) but showed lower sensitivity (86% for CT < 30) than the RT-LAMP assay. In addition, we developed a multiplexed sequencing protocol (LAMP-sequencing) as a diagnostic validation procedure to detect and record the outcome of RT-LAMP reactions.