4.4 Article

Fluorescence-based Neuraminidase Inhibition Assay to Assess the Susceptibility of Influenza Viruses to The Neuraminidase Inhibitor Class of Antivirals

Journal

JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
Volume -, Issue 122, Pages -

Publisher

JOURNAL OF VISUALIZED EXPERIMENTS
DOI: 10.3791/55570

Keywords

Immunology; Issue 122; Fluorescence; neuraminidase; resistance; influenza; MUNANA; antiviral; inhibitor; phenotypic assay; oseltamivir; zanamivir

Funding

  1. Australian Government Department of Health

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The neuraminidase (NA) inhibitors are the only class of antivirals approved for the treatment and prophylaxis of influenza that are effective against currently circulating strains. In addition to their use in treating seasonal influenza, the NA inhibitors have been stockpiled by a number of countries for use in the event of a pandemic. It is therefore important to monitor the susceptibility of circulating influenza viruses to this class of antivirals. There are different types of assays that can be used to assess the susceptibility of influenza viruses to the NA inhibitors, but the enzyme inhibition assays using either a fluorescent substrate or a chemiluminescent substrate are the most widely used and recommended. This protocol describes the use of a fluorescence-based assay to assess influenza virus susceptibility to NA inhibitors. The assay is based on the NA enzyme cleaving the 2'-(4-Methylumbelliferyl)-alpha-D-N-acetylneuraminic acid (MUNANA) substrate to release the fluorescent product 4-methylumbelliferone (4-MU). Therefore, the inhibitory effect of an NA inhibitor on the influenza virus NA is determined based on the concentration of the NA inhibitor that is required to reduce 50% of the NA activity, given as an IC50 value.

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