Sample-to-answer COVID-19 nucleic acid testing using a low-cost centrifugal microfluidic platform with bead-based signal enhancement and smartphone read-out

• Published in: Lab on a Chip Vol. 21; no. 15; pp. 2932 - 2944
• Main Authors: Soares, Ruben R. G, Akhtar, Ahmad S, Pinto, Inês F, Lapins, Noa, Barrett, Donal, Sandh, Gustaf, Yin, Xiushan, Pelechano, Vicent, Russom, Aman
• Format: Journal Article Web Resource
• Language: English
• Published: Cambridge Royal Society of Chemistry 2021
• Subjects: Alkanolamines; Beads; Centrifugal microfluidic platform; Centrifugation; Coronaviruses; Cost effectiveness; Costs; COVID-19; COVID-19 Testing; Deactivation; Detection limits; Diagnosis; Dimers; Disease transmission; Diseases; Enhanced fluorescence; Fluorescence; genetics; human; Humans; Income; Infektionsmedicin; Klinisk medicin; Loop mediated isothermal amplifications; Medicin och hälsovetenskap; Medicinsk bioteknologi; Medicinsk bioteknologi (inriktn. mot cellbiologi (inkl. stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci); Microfluidics; molecular diagnosis; Molecular Diagnostic Techniques; nucleic acid amplification; Nucleic Acid Amplification Techniques; Nucleic acids; Public health; Reaction volume; Ribonucleic acid; RNA; Sample collection; SARS-CoV-2; sensitivity and specificity; Severe acute respiratory syndrome coronavirus 2; Signal enhancement; Smart-phone cameras; smartphone; Smartphones; System effectiveness; Viral; Viral diseases; virus RNA
• ISSN: 1473-0197; 1473-0189; 1473-0189
• DOI: 10.1039/d1lc00266j

With its origin estimated around December 2019 in Wuhan, China, the ongoing SARS-CoV-2 pandemic is a major global health challenge. The demand for scalable, rapid and sensitive viral diagnostics is thus particularly pressing at present to help contain the rapid spread of infection and prevent overwhelming the capacity of health systems. While high-income countries have managed to rapidly expand diagnostic capacities, such is not the case in resource-limited settings of low- to medium-income countries. Aiming at developing cost-effective viral load detection systems for point-of-care COVID-19 diagnostics in resource-limited and resource-rich settings alike, we report the development of an integrated modular centrifugal microfluidic platform to perform loop-mediated isothermal amplification (LAMP) of viral RNA directly from heat-inactivated nasopharyngeal swab samples. The discs were pre-packed with dried n -benzyl- n -methylethanolamine modified agarose beads used to selectively remove primer dimers, inactivate the reaction post-amplification and allowing enhanced fluorescence detection via a smartphone camera. Sample-to-answer analysis within 1 hour from sample collection and a detection limit of approximately 100 RNA copies in 10 μL reaction volume were achieved. The platform was validated with a panel of 162 nasopharyngeal swab samples collected from patients with COVID-19 symptoms, providing a sensitivity of 96.6% (82.2-99.9%, 95% CI) for samples with Ct values below 26 and a specificity of 100% (90-100%, 95% CI), thus being fit-for-purpose to diagnose patients with a high risk of viral transmission. These results show significant promise towards bringing routine point-of-care COVID-19 diagnostics to resource-limited settings. LAMP-based platform for SARS-CoV-2 RNA detection incorporating beads to remove primer-dimers, enhance fluorescent signal and stop the reaction after amplification.