Clinical Performance of Reverse Transcription Loop-mediated Isothermal Amplification COVID-19 Assay on Gold-nanoparticle-modified Screen-printed Carbon Electrode Using Differential Pulse Voltammetry

Abstract

The World Health Organization (WHO) has recommended real-time reverse transcription polymerase chain reaction (RT-PCR) as the gold standard for coronavirus disease detection. In this study, we aim to validate the clinical performance of reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay on a gold-nanoparticle-modified screen-printed carbon electrode (AuNP/SPCE) using differential pulse voltammetry (DPV) and to compare it with real-time RT-PCR. The electrodeposited AuNP on SPCE was quasi-spherical with a size of ±500 nm. The developed RT-LAMP primer was designed from the GenBank database using the NCBI Multiple Alignment tools and Jalview software. Nasopharyngeal clinical samples were obtained from suspected COVID-19 patients (n = 148). The RT-LAMP products were dropped on the modified AuNP/SPCE under DPV setting, which resulted in current change (∆I) responses. The positive and negative samples produced significantly different ∆I signals with a p-value <0.0001 at a 95% confidence interval using Student’s t-test. The RT-LAMP assay using Au/SPCE exhibited a 30 s response time per analysis. The clinical sensitivity and specificity obtained were 79.7 and 85.1%, respectively, with a detection limit of 0.4 copies µl−1. Hence, this proposed method is suitable for COVID-19 RNA detection in resource-limited settings.