Year: 2024
Type: conference-paper
Abstract: This paper reports a digital focus assay (dFA) that addresses several limitations of conventional assays for quantifying infectious influenza virus.The preliminary results using this platform were reported at µTAS 2023 (poster) and AIChE Annual Meeting 2023 1 .This work builds on that through the i) Use of an aqueous maintenance media (MM) in addition to a viscous, microcrystalline cellulose (MCC)-based overlay medium, which simplified the device operation ii) Automation of the image analysis using a custom Python code iii) Verification of the dFA viral titers by comparing against a plaque assay performed in parallel iv) Demonstration of running clinical samples on the dFA platform with comparisons to the corresponding dPCR titers.Despite being rapid and sensitive, assays like the PCR, ELISA cannot distinguish between viable, infectious virus and non-viable virus, necessitating cell-culture based assays.Traditional plaque assays take multiple days to provide a readout, are not very sensitive, and do not work for non-lytic viruses.Traditional focus forming assays (FFA) despite being the fastest and the most sensitive of cell-culture based assays, suffer from many drawbacks such as high costs due to antibodies needed for immunostaining, laborious manual counting of the foci, and a narrow endpoint time window.The dFA matches the speed of the FFA while overcoming these drawbacks through significantly reduced reagent consumption for lower assay costs, automated image analysis of the readout for convenient operation and elimination of subjectivity in interpreting the results, and introduction of flexible endpoint timing enabled by the digital architecture (Fig. 1).
DOI: 10.70477/rehz5168
Institution: University of Maryland, College Park
Language: English
Subfield: Biomedical Engineering
Field: Engineering
Domain: Physical Sciences
Open Access status: gold
OpenAlex ID: W4408828329