“An Open One-Step RT-qPCR for SARS-CoV-2 detection”
Abstract
The COVID-19 pandemic has resulted in millions of deaths globally, and while several diagnostic systems were proposed, real-time reverse transcription polymerase chain reaction (RT-PCR) remains the gold standard. However, diagnostic reagents, including enzymes used in RT-PCR, are subject to centralized production models and intellectual property restrictions, which present a challenge for less developed countries. With the aim of generating a standardized One-Step open RT-qPCR protocol to detect SARS-CoV-2 RNA in clinical samples, we purified and tested recombinant enzymes and a non-proprietary buffer. The protocol utilized M-MLV RT and Taq DNA pol enzymes to perform a Taqman probe-based assay. Synthetic RNA samples were used to validate the One-Step RT-qPCR components, demonstrating sensitivity comparable to a commercial kit routinely employed in clinical settings for patient diagnosis. Further evaluation on 40 clinical samples (20 positive and 20 negative) confirmed its comparable diagnostic accuracy. This study represents a proof of concept for an open approach to developing diagnostic kits for viral infections and diseases, which could provide a cost-effective and accessible solution for less developed countries.
Huge congratulations to @ArielCerda, @mairarivera, Grace Armijo, Catalina Ibarra-Henriquez, Javiera Reyes, Paula Blázquez-Sánchez, Javiera Avilés, @AnibalArc, Aldo Seguel, @A.Brown, Yesseny Vásquez, Marcelo Cortez-San Martín, Francisco A. Cubillos, Patricia García, Marcela Ferres, @cesarramirez, @FernanFederici, and Rodrigo A. Gutiérrez for this multi-institutional body of work!
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