Laboratory methodologies for testing the antiviral susceptibility of influenza viruses:
Neuraminidase inhibitor (NAI)

Phenotyping: neuraminidase inhibition assays

Two types of neuraminidase (NA) inhibition assays are commonly used for determining influenza susceptibility to the NA inhibitor (NAI) antivirals: fluorescence-based (FL) and chemiluminescence-based (CL) assays.

Both types of assays have advantages and disadvantages (see summary below). For practical reasons, the FL assays are more cost-effective and applicable, in particular for National Influenza Centres (NICs) in the process of establishing NA inhibition assays.

NICs are encouraged to contact any of the WHO Collaborating Centres in GISRS when establishing NA inhibition assays, to ensure continuous support, including provision/updates of protocols, control viruses, etc.

Protocols available for NA inhibition FL assays are:

  • an ‘in-house’ FL method (see the WHO Manual for the laboratory diagnosis and virological surveillance of influenza);
  • commercially available kits (see summary below).

Software for calculating IC50 values from NA inhibition assay data

Various software programs are available for the calculation of IC50 values from NA enzyme inhibition assay data. Variations in IC50 values may occur as a result of differences in the software programs.

  • GraphPad – commercially available software:
  • GraFit – commercially available software:
  • JASPR – freely available software developed by the Centers for Disease Control and Prevention (CDC), Atlanta, United States of America. To request the software please contact FLUANTIVIRAL@CDC.GOV or contact Dr. Larisa Gubareva ( for any further assistance.
  • PHE point-to-point – freely available analysis developed by the Health Protection Agency, London, United Kingdom of Great Britain and Northern Ireland. Please contact Dr. Angie Lackenby ( to request the template and protocol.