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 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, spike-expressing 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 the infectious disease.