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High-throughput viral microneutralization method for feline coronavirus using image cytometry.
Pearson, Morgan; LaVoy, Alora; Chan, Leo Li-Ying; Dean, Gregg A.
  • Pearson M; Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO, 80523, United States.
  • LaVoy A; Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO, 80523, United States.
  • Chan LL; Department of Advanced Technology R&D, Nexcelom Bioscience LLC, Lawrence, MA, 01843, United States. Electronic address: lchan@nexcelom.com.
  • Dean GA; Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO, 80523, United States.
J Virol Methods ; 286: 113979, 2020 12.
Article in English | MEDLINE | ID: covidwho-786045
ABSTRACT
Feline coronaviruses (FCoV) are members of the alphacoronavirus genus that are further characterized by serotype (types I and II) based on the antigenicity of the spike (S) protein and by pathotype based on the associated clinical conditions. Feline enteric coronaviruses (FECV) are associated with the vast majority of infections and are typically asymptomatic. Within individual animals, FECV can mutate and cause a severe and usually fatal disease called feline infectious peritonitis (FIP), the leading infectious cause of death in domestic cat populations. There are no approved antiviral drugs or recommended vaccines to treat or prevent FCoV infection. The plaque reduction neutralization test (PRNT) traditionally employed to assess immune responses and to screen therapeutic and vaccine candidates is time-consuming, low-throughput, and typically requires 2-3 days for the formation and manual counting of cytolytic plaques. Host cells are capable of carrying heavy viral burden in the absence of visible cytolytic effects, thereby reducing the sensitivity of the assay. In addition, operator-to-operator variation can generate uncertainty in the results and digital records are not automatically created. To address these challenges we developed a novel high-throughput viral microneutralization assay, with quantification of virus-infected cells performed in a plate-based image cytometer. Host cell seeding density, microplate surface coating, virus concentration and incubation time, wash buffer and fluorescent labeling were optimized. Subsequently, this FCoV viral neutralization assay was used to explore immune correlates of protection using plasma from naturally FECV-infected cats. We demonstrate that the high-throughput viral neutralization assay using the Celigo Image Cytometer provides a robust and efficient method for the rapid screening of therapeutic antibodies, antiviral compounds, and vaccines. This method can be applied to various viral infectious diseases to accelerate vaccine and antiviral drug discovery and development.
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Full text: Available Collection: International databases Database: MEDLINE Main subject: Neutralization Tests / Coronavirus Infections / Coronavirus, Feline / Image Cytometry / High-Throughput Screening Assays Type of study: Diagnostic study / Prognostic study Topics: Vaccines Limits: Animals Language: English Journal: J Virol Methods Year: 2020 Document Type: Article Affiliation country: J.jviromet.2020.113979

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Neutralization Tests / Coronavirus Infections / Coronavirus, Feline / Image Cytometry / High-Throughput Screening Assays Type of study: Diagnostic study / Prognostic study Topics: Vaccines Limits: Animals Language: English Journal: J Virol Methods Year: 2020 Document Type: Article Affiliation country: J.jviromet.2020.113979