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Viruses 2012, 4(12), 3511-3530; doi:10.3390/v4123511

Standardization of the Filovirus Plaque Assay for Use in Preclinical Studies

Received: 12 October 2012 / Revised: 10 November 2012 / Accepted: 20 November 2012 / Published: 6 December 2012
(This article belongs to the Special Issue Advances in Filovirus Research 2012)
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The filovirus plaque assay serves as the assay of choice to measure infectious virus in a cell culture, blood, or homogenized tissue sample. It has been in use for more than 30 years and is the generally accepted assay used to titrate virus in samples from animals treated with a potential antiviral therapeutic or vaccine. As these animal studies are required for the development of vaccines and therapeutics under the FDA Animal Rule, it is essential to have a standardized assay to compare their efficacies against the various filoviruses. Here, we present an evaluation of the conditions under which the filovirus plaque assay performs best for the Ebola virus Kikwit variant and the Angola variant of Marburg virus. The indicator cell type and source, inoculum volumes, length of incubation and general features of filovirus biology as visualized in the assay are addressed in terms of the impact on the sample viral titer calculations. These optimization studies have resulted in a plaque assay protocol which can be used for preclinical studies, and as a standardized protocol for use across institutions, to aid in data comparison. This protocol will be validated for use in GLP studies supporting advanced development of filovirus therapeutics and vaccines.
Keywords: plaque assay; filovirus; Ebola; ebolavirus; marburgvirus; Marburg virus; Vero plaque assay; filovirus; Ebola; ebolavirus; marburgvirus; Marburg virus; Vero
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Shurtleff, A.C.; Biggins, J.E.; Keeney, A.E.; Zumbrun, E.E.; Bloomfield, H.A.; Kuehne, A.; Audet, J.L.; Alfson, K.J.; Griffiths, A.; Olinger, G.G.; Bavari, S. Standardization of the Filovirus Plaque Assay for Use in Preclinical Studies. Viruses 2012, 4, 3511-3530.

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