A <u>S</u>imple, <u>A</u>ffordable, <u>R</u>apid, <u>S</u>tabilized, <u>Co</u>lorimetric, <u>V</u>ersatile RT-LAMP Assay to Detect SARS-CoV-2
Juan García-Bernalt Diego,
Pedro Fernández-Soto,
Marta Domínguez-Gil,
Moncef Belhassen-García,
Juan Luis Muñoz Bellido,
Antonio Muro
Affiliations
Juan García-Bernalt Diego
Infectious and Tropical Diseases Research Group (e-INTRO), Biomedical Research Institute of Salamanca-Research Centre for Tropical Diseases at the University of Salamanca (IBSAL-CIETUS), Faculty of Pharmacy, University of Salamanca, 37007 Salamanca, Spain
Pedro Fernández-Soto
Infectious and Tropical Diseases Research Group (e-INTRO), Biomedical Research Institute of Salamanca-Research Centre for Tropical Diseases at the University of Salamanca (IBSAL-CIETUS), Faculty of Pharmacy, University of Salamanca, 37007 Salamanca, Spain
Marta Domínguez-Gil
Microbiology Service, Hospital Universitario Río Hortega, 47012 Valladolid, Spain
Moncef Belhassen-García
Infectious and Tropical Diseases Research Group (e-INTRO), Biomedical Research Institute of Salamanca-Research Centre for Tropical Diseases at the University of Salamanca (IBSAL-CIETUS), Faculty of Pharmacy, University of Salamanca, 37007 Salamanca, Spain
Juan Luis Muñoz Bellido
Microbiology and Parasitology Service, Complejo Asistencial Universitario de Salamanca, University of Salamanca, 37007 Salamanca, Spain
Antonio Muro
Infectious and Tropical Diseases Research Group (e-INTRO), Biomedical Research Institute of Salamanca-Research Centre for Tropical Diseases at the University of Salamanca (IBSAL-CIETUS), Faculty of Pharmacy, University of Salamanca, 37007 Salamanca, Spain
The SARS-CoV-2 pandemic has forced all countries worldwide to rapidly develop and implement widespread testing to control and manage the Coronavirus Disease 2019 (COVID-19). reverse-transcription (RT)-qPCR is the gold standard molecular diagnostic method for COVID-19, mostly in automated testing platforms. These systems are accurate and effective, but also costly, time-consuming, high-technological, infrastructure-dependent, and currently suffer from commercial reagent supply shortages. The reverse-transcription loop-mediated isothermal amplification (RT-LAMP) can be used as an alternative testing method. Here, we present a novel versatile (real-time and colorimetric) RT-LAMP for the simple (one-step), affordable (~1.7 €/sample), and rapid detection of SARS-CoV-2 targeting both ORF1ab and N genes of the novel virus genome. We demonstrate the assay on RT-qPCR-positive clinical samples, obtaining most positive results under 25 min. In addition, a novel 30-min one-step drying protocol has been developed to stabilize the RT-LAMP reaction mixtures, allowing them to be stored at room temperature functionally for up to two months, as predicted by the Q10. This Dry-RT-LAMP methodology is suitable for potentially ready-to-use COVID-19 diagnosis. After further testing and validation, it could be easily applied both in developed and in low-income countries yielding rapid and reliable results.
