A Magnetic Modulation Biosensing-Based Molecular Assay for Rapid and Highly Sensitive Clinical Diagnosis of Coronavirus Disease 2019 (COVID-19)

Rapid and sensitive detection of human pathogens, such as the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), is an urgent and challenging task for clinical laboratories. Currently, the gold standard test for SARS-CoV-2–specific RNA is based on quantitative RT-PCR (RT-qPCR), which relies on target amplification by Taq polymerase and uses a fluorescent resonance energy transfer–based hydrolysis probe. Although this method is accurate and specific, it is also time consuming. To rapidly detect the presence of the viral RNA in clinical samples, we describe a new molecular assay that combines a highly sensitive magnetic modulation biosensing (MMB) system, rapid thermal cycling, and a modified double-quenched hydrolysis probe. Using in vitro transcribed SARS-CoV-2 RNA targets spiked in PCR-grade water, we found that the calculated limit of detection of the MMB-based molecular assay was 1.6 copies per reaction. Testing 309 RNA extracts from 170 confirmed RT-qPCR SARS-CoV-2–negative individuals (30 of whom were positive to other respiratory viruses) and 139 RT-qPCR SARS-CoV-2–positive patients (CT ≤ 42) resulted in 97.8% sensitivity, 100% specificity, and 0% cross-reactivity. The total turnaround time of the MMB-based assay is 30 minutes, which is three to four times faster than a standard RT-qPCR. By adjusting the primers and the probe set, the platform can be easily adapted to detect most of the pathogens that are currently being diagnosed by RT-qPCR.