Quantitation of flaviviruses by fluorescent focus assay

Journal of Virological Methods
Anne F PayneLaura D Kramer

Abstract

An indirect immunofluorescence assay for quantitation of flaviviruses was developed as an alternative to the standard plaque assay. The assay was validated with West Nile virus (WNV), St. Louis encephalitis virus (SLEV), and Dengue virus (DENV) types 1-4. Vero cells were plated in 8-well chamber slides, and infected with 10-fold serial dilutions of virus. About 1-3 days after infection, cells were fixed, incubated with specific monoclonal antibody, and stained with a secondary antibody labeled with a fluorescent tag. Fluorescent foci of infection were observed and counted using a fluorescence microscope, and viral titers were calculated as fluorescent focus units (FFU) per ml. The optimal time for performing the fluorescent focus assay (FFA) on Vero cells was 24 h for WNV, and 48 h for SLEV and the four DENV serotypes. In contrast, the time required to complete a standard Vero cell plaque assay for these viruses range from 3 days for WNV to 11 days for DENV-1. Thus, the FFA method of virus titration is useful for viruses whose plaques develop slowly. In addition, these viruses can be quantitated by FFA on a mosquito cell line (C6/36), which does not support plaque formation. The FFA for flaviviruses was validated for accuracy, ...Continue Reading

Citations

May 24, 2011·Analytical Chemistry·Gleb G MironovMaxim V Berezovski
Oct 31, 2009·Vector Borne and Zoonotic Diseases·A Marm KilpatrickLaura D Kramer
Aug 27, 2011·Vector Borne and Zoonotic Diseases·Brittany L DodsonJason L Rasgon
Jul 10, 2009·Journal of Virology·Dipankar BhattacharyaRob Striker
May 1, 2012·BMC Evolutionary Biology·Alexander T CiotaLaura D Kramer
Jul 11, 2009·BMC Evolutionary Biology·Louis LambrechtsThomas W Scott
Jun 11, 2011·Virology Journal·Katherine M SmithRaquel Hernandez
Oct 3, 2012·PLoS Neglected Tropical Diseases·Agnieszka M SzemielRichard M Elliott
Apr 5, 2013·PLoS Neglected Tropical Diseases·Marcus S C BlagroveSteven P Sinkins
May 3, 2013·PLoS Neglected Tropical Diseases·Lauren B CarringtonThomas W Scott
Dec 12, 2012·Viruses·Amy C ShurtleffUNKNOWN Filovirus Animal Nonclinical Group (FANG) Assay Working Group
Sep 3, 2010·The American Journal of Tropical Medicine and Hygiene·A Marm KilpatrickLaura D Kramer
Sep 8, 2011·The American Journal of Tropical Medicine and Hygiene·Gillian EastwoodAndrew A Cunningham
Jul 8, 2011·The American Journal of Tropical Medicine and Hygiene·Laura D KramerGraham Mackereth
Jul 18, 2012·The American Journal of Tropical Medicine and Hygiene·Jorge E OsorioErik Hofmeister
Feb 27, 2013·The American Journal of Tropical Medicine and Hygiene·Lauren B CarringtonThomas W Scott
Feb 3, 2009·Virus Research·Dipankar BhattacharyaRob Striker
Oct 23, 2010·Cellular Microbiology·Noelia López-Montero, Cristina Risco
Dec 18, 2013·Biotechnology and Bioengineering·Adam SwickJohn Yin
Jun 19, 2008·The Journal of General Virology·Alexander T CiotaLaura D Kramer
Mar 20, 2014·The Journal of General Virology·Alexander T CiotaLaura D Kramer
Aug 21, 2013·Biological Conservation·A Marm KilpatrickLaura D Kramer
Feb 14, 2012·Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases·Alexander T CiotaLaura D Kramer

❮ Previous
Next ❯

Related Concepts

Related Feeds

Aminoglycosides

Aminoglycoside is a medicinal and bacteriologic category of traditional Gram-negative antibacterial medications that inhibit protein synthesis and contain as a portion of the molecule an amino-modified glycoside. Discover the latest research on aminoglycoside here.

Aminoglycosides (ASM)

Aminoglycoside is a medicinal and bacteriologic category of traditional Gram-negative antibacterial medications that inhibit protein synthesis and contain as a portion of the molecule an amino-modified glycoside. Discover the latest research on aminoglycoside here.