An Optimized High-Throughput Immuno-Plaque Assay for SARS-CoV-2

• Main Authors: Alberto A. Amarilla, Naphak Modhiran, Yin Xiang Setoh, Nias Y. G. Peng, Julian D. J. Sng, Benjamin Liang, Christopher L. D. McMillan, Morgan E. Freney, Stacey T. M. Cheung, Keith J. Chappell, Alexander A. Khromykh, Paul R. Young, Daniel Watterson
• Format: Article
• Language: English
• Published: Frontiers Media S.A. 2021-02-01
• Series: Frontiers in Microbiology
• Subjects: coronaviruses; SARS-CoV-2; immuno-plaque assay (iPA); viral quantification
• Online Access: https://www.frontiersin.org/articles/10.3389/fmicb.2021.625136/full

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has been identified as the causative agent of coronavirus disease 2019 and is capable of human-to-human transmission and rapid global spread. The rapid emergence and global spread of SARS-CoV-2 has encouraged the establishment of a rapid, sensitive, and reliable viral detection and quantification methodology. Here, we present an alternative assay, termed immuno-plaque assay (iPA), which utilizes a combination of plaque assay and immunofluorescence techniques. We have extensively optimized the conditions for SARS-CoV-2 infection and demonstrated the great flexibility of iPA detection using several antibodies and dual-probing with two distinct epitope-specific antibodies. In addition, we showed that iPA could be utilized for ultra-high-throughput viral titration and neutralization assay within 24 h and is amenable to a 384-well format. These advantages will significantly accelerate SARS-CoV-2 research outcomes during this pandemic period.