Multiparametric Integrated Molecular Detection of SARS-CoV-2 from Biofluids by Adapting Single Extracellular Vesicle Characterization Technologies

Background: The purpose of this study was to develop a biochip assay capable of simultaneously detecting SARS-CoV-2 protein and RNA from biofluids with single-virus resolution. COVID-19, caused by the SARS-CoV-2 virus, had rapidly progressed into a global pandemic, necessitating more efficient and precise diagnostic tools beyond conventional nucleic acid, serological, and antigen assays. Materials/Methods: The study designed a biochip assay that utilized surface markers to concurrently detect and quantify viral proteins and RNA. Signal amplification was achieved by targeting multiple regions of the viral RNA, including nucleocapsid N1 and N2, while tyramide signal amplification (TSA) enhanced detection of viral surface proteins. The assay was applied to various biofluids, including nasopharyngeal swabs and saliva samples collected from COVID-19 patients, to demonstrate its clinical applicability. Outcome/Impact: The study demonstrated that the biochip assay successfully detected SARS-CoV-2 protein and RNA simultaneously from patient samples, confirming its clinical feasibility. The intended outcome was to establish a sensitive, multiplexed diagnostic platform for rapid and accurate COVID-19 detection across diverse sample types.