title: A quantitative comet infection assay for influenza virus.
abstract: The virus comet assay is a cell-based virulence assay used to evaluate an antiviral drug or antibody against a target virus. The comet assay differs from the plaque assay in allowing spontaneous flows in 6-well plates to spread virus. When implemented quantitatively the comet assay has been shown to have an order-of-magnitude greater sensitivity to antivirals than the plaque assay. In this study, a quantitative comet assay for influenza virus is demonstrated, and is shown to have a 13-fold increase in sensitivity to ribavirin. AX4 cells (MDCK cells with increased surface concentration of α2-6 sialic acid, the influenza virus receptor) have reduced the comet size variability relative to MDCK cells, making them a better host cell for use in this assay. Because of enhanced antiviral sensitivity in flow-based assays, less drug is required, which could lead to lower reagent costs, reduced cytotoxicity, and fewer false-negative drug screen results. The comet assay also serves as a readout of flow conditions in the well. Observations from comets formed at varying humidity levels indicate a role for evaporation in the mechanism of spontaneous fluid flow in wells.
authors.affiliation: Department of Chemical and Biological Engineering, Systems Biology Theme, Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA. smlindsay@wisc.edu
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Lindsay
authors.forename: Andrea
authors.initials: A
authors.lastname: Timm
authors.forename: John
authors.initials: J
authors.lastname: Yin
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology; D000914:Antibodies, Viral / Q000276:immunology; D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D004285:Dogs; D006813:Humidity; D009975:Orthomyxoviridae / Q000187:drug effects* / Q000276:immunology / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D019562:Viral Load / Q000191:economics / Q000379:methods*; D010948:Viral Plaque Assay / Q000379:methods
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D000998:Antiviral Agents