The Development of Aptamer-Functionalized Streptavidin Magnetic Particles for Sample Purification of SARS-CoV-2 Coupled with Rapid Plasmonic RT-qPCR to Improve Detection Sensitivity

Abstract

Despite having endured numerous pandemics throughout history and contemporary technological advancements, the world was still unprepared for the emergence of the SARS-CoV-2 pathogen.Prior to the development and distribution of effective vaccines, health officials relied heavily on diagnostics to contain the spread of the virus. Reverse transcription-polymerase chain reaction (RT-PCR) is employed for the diagnosing SARS-CoV-2 due to its exceptional sensitivity and specificity. However, clinical sample preparation techniques for SARS-CoV-2 may have limited detection sensitivity due to the large volumes of viral transport medium in which nasopharyngeal swabs are stored, and where only a fraction of the sample was used for analysis. Consequently, we have developed a sample processing method that improves the sensitivity and specificity of SARS-CoV-2 diagnostics by capturing viral particles using aptamers specific to the S-protein functionalized on magnetic particles ("Apta-beads") and thereby concentrating the analyte. The eluted viral particles are then directly amplified on the Kimera P-IV Point-of-Care Plasmonic qPCR platform. With our protocols, we have been able to capture various forms of SARS-CoV-2 analytes, including recombinant spike protein from both Wuhan and Omicron strains, spikeexpressing pseudoviral particles, and SARS-CoV-2 viruses. Furthermore, the capture protocol can be performed in as little as 2 minutes, with the added effect of removing unwanted inhibitory components to PCR reactions. In conclusion, our "Apta-bead" sample purification, combined with the rapid Plasmonic PCR analysis, can play a critical role in limiting sample-to-result turnover time, and thereby mitigate the spread of infectious disease.