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Solid-Phase Assays of Receptor-Binding Specificity

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Influenza Virus

Part of the book series: Methods in Molecular Biology ((MIMB,volume 865))

Abstract

Influenza virus attachment to sialic acid-containing molecules on the cell surface initiates the infection. The spectrum of functional receptors on target cells and decoy receptors on cells and epithelial mucus varies substantially between animal species leading to variations in the receptor-binding specificity of viruses circulating in these species. Analysis of the receptor specificity of different animal and human influenza viruses can give insight into factors and mechanisms that determine viral host range, tissue and cell tropism, replication efficiency, and pathogenesis. Knowledge of viral receptor specificity may also be useful for the development of more efficient influenza vaccines and anti-influenza drugs.

A majority of known receptor specificity assays measure influenza virus binding to sialic acid-containing natural and synthetic compounds (receptor analogues). Here, we describe protocols of two solid-phase enzyme-linked receptor-binding assays which are technically similar to standard ELISA. Each assay determines binding of the virus immobilized in the wells of 96-well plate to receptor analogues in solution. In the direct binding assay, the virus binds to either synthetic biotinylated sialylglycopolymers or to peroxidase-labeled sialylglycoprotein fetuin (Fet-HRP); the apparent association constants of the virus–receptor complexes are calculated from the Scatchard plots of the binding data. In the fetuin-binding inhibition assay, the virus is incubated with a mixture of unlabeled receptor analogue and standard preparation of Fet-HRP; the association constant for analogue is calculated based on the level of its competition with Fet-HRP.

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Acknowledgments

We thank Prof. Nicolai Bovin and members of the Carbohydrate Laboratory at the Institute of Bio-organic Chemistry, Moscow for providing sialylglycopolymers and for long-term collaboration. We are grateful to Prof. Hans-Dieter Klenk for his interest and stimulating discussions. We acknowledge recent financial support of our own work from the Deutsche Forschungsgemeinschaft (SFB 593), the European Commission projects ESNIP-2, FLUPATH, FLUINNATE, and FLUPIG, and from the Russian Foundation for Basic Research (grant 08-04-00691).

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Authors and Affiliations

  1. Institute of Virology, Philipps University, Marburg, Germany

    Mikhail N. Matrosovich

  2. M.P. Chumakov Institute of Poliomyelitis and Viral Encephalitides, Russian Academy of Medical Sciences, Moscow, Russian Federation

    Alexandra S. Gambaryan

Authors
  1. Mikhail N. Matrosovich
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  2. Alexandra S. Gambaryan
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Correspondence to Mikhail N. Matrosovich .

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Editors and Affiliations

  1. School of Veterinary Medicine, Dept. Pathobiological Science, University of Wisconsin-Madison, Science Drive 575, Madison, 53711, Wisconsin, USA

    Yoshihiro Kawaoka

  2. Influenza Research Institute (IRI), Dept. Pathobiological Science, University of Wisconsin-Madison, Science Drive 575, Madison, 53711, Wisconsin, USA

    Gabriele Neumann

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Matrosovich, M.N., Gambaryan, A.S. (2012). Solid-Phase Assays of Receptor-Binding Specificity. In: Kawaoka, Y., Neumann, G. (eds) Influenza Virus. Methods in Molecular Biology, vol 865. Humana Press. https://doi.org/10.1007/978-1-61779-621-0_5

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  • DOI: https://doi.org/10.1007/978-1-61779-621-0_5

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-620-3

  • Online ISBN: 978-1-61779-621-0

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