A screening assay for neuraminidase inhibitors using neuraminidases N1 and N3 from a baculovirus expression system

Abstract

Context: Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibitors.

Objective: Validate the use of recombinant neuraminidase expressed in baculovirus located on the viral surface capsule to develop a neuraminidase inhibitor screening assay.

Materials and methods: Recombinant baculovirus particles displaying neuraminidase N1 and N3 were used as enzyme sources. The assay set-up required the use of 2′-(4-methylumbelliferyl)-α-D-acetyl neuraminic acid as substrate and oseltamivir carboxylate as benchmark inhibitor.

Results: The assay was set up in a standard 96-well plate. The within- and between-assay coefficients of variation were, on average, less than 10%. The 50% inhibitory concentration values of the inhibitor were in good agreement with those determined by independent kinetic experiments.

Discussion and conclusions: The assay showed satisfactory within- and between-assay repeatability. The obtained results suggest that recombinant baculovirus expressing neuraminidase located on the virus membrane capsule can be used to set up affordable and reliable neuraminidase inhibitors screening assays.

Keywords::

• Avian influenza
• neuraminidase inhibitor
• bioassay
• baculovirus

Acknowledgments

J. K. would like to thank Chulalongkorn University and The Embassy of Italy in Thailand for supporting her Ph.D. scholarship.

Declaration of interest

This work was supported by the Joint Project between the International Centre for Science and High Technology (UNIDO) Trieste, Italy and the Office of the Higher Education Commission, Bangkok, Thailand.