Abstract
Here we describe methods for the in vivo study of antiviral NK cell responses using the mouse Orthopoxvirus ectromelia virus as a model, the agent of mousepox. The methods include those specific for the preparation and use of ectromelia virus such as the production of virus stocks in tissue culture and in live mice, the purification of virus stocks, the titration of virus stocks and virus loads in organs, and the infection of mice. The chapter also includes methods for the specific study of NK cell responses in infected mice such as the preparation of organs (lymph nodes, spleen, and liver) for analysis, the study of NK cell responses by flow cytometry, the adoptive transfer of NK cells, the measurement of NK cell cytolytic activity ex vivo and in vivo, and the determination of NK cell proliferation by bromodeoxyuridine loading or by dilution of carboxyfluorescein diacetate succinimidyl ester (CFSE).
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References
French, A. R., and Yokoyama, W. M. (2003) Natural killer cells and viral infections. Curr Opin Immunol 15, 45–51.
Biron, C. A., Nguyen, K. B., Pien, G. C., Cousens, L. P., and Salazar-Mather, T. P. (1999) Natural killer cells in antiviral defense: function and regulation by innate cytokines. Annu Rev Immunol 17, 189–220.
Arase, H., Mocarski, E. S., Campbell, A. E., Hill, A. B., and Lanier, L. L. (2002) Direct recognition of cytomegalovirus by activating and inhibitory NK cell receptors. Science 296, 1323–6.
Daniels, K. A., Devora, G., Lai, W. C., O'Donnell, C. L., Bennett, M., and Welsh, R. M. (2001) Murine cytomegalovirus is regulated by a discrete subset of natural killer cells reactive with monoclonal antibody to Ly49H. J Exp Med 194, 29–44.
Lee, S. H., Girard, S., Macina, D., Busa, M., Zafer, A., Belouchi, A., Gros, P., and Vidal, S. M. (2001) Susceptibility to mouse cytomegalovirus is associated with deletion of an activating natural killer cell receptor of the C-type lectin superfamily. Nat Genet 28, 42–5.
Jacoby, R. O., Bhatt, P. N., and Brownstein, D. G. (1989) Evidence that NK cells and interferon are required for genetic resistance to lethal infection with ectromelia virus. Arch Virol 108, 49–58.
Parker, A. K., Parker, S., Yokoyama, W. M., Corbett, J. A., and Buller, R. M. (2007) Induction of natural killer cell responses by ectromelia virus controls infection. J Virol 81, 4070–9.
Fang, M., Lanier, L. L., and Sigal, L. J. (2008) A role for NKG2D in NK cell-mediated resistance to poxvirus disease. PLoS Pathog 4, e30.
Fenner, F.,Henderson, D. A., Arita, I., Jezek, Z., Ladnyi, D., and Organization, W. H. (1988) Smallpox and its eradication, World Health Organization, Geneva. Chapter 1, 2–68.
Delano, M. L., and Brownstein, D. G. (1995) Innate resistance to lethal mousepox is genetically linked to the NK gene complex on chromosome 6 and correlates with early restriction of virus replication by cells with an NK phenotype. J Virol 69, 5875–7.
Earl, P. L., Moss, B., Wyatt, L. S., and Carroll, M. W. (2001) Generation of recombinant vaccinia viruses. Curr Protoc Mol Biol Chapter 16, Unit 16 17.
Scalzo, A. A., Farrell, H. E., and Karupiah, G. (2000) Techniques for studying murine natural killer cells in defense against viral infection. Methods Mol Biol 121, 163–77.
Lyons, A. B., and Parish, C. R. (1994) Determination of lymphocyte division by flow cytometry. J Immunol Methods 171, 131–7.
Barber, D. L., Wherry, E. J., and Ahmed, R. (2003) Cutting edge: rapid in vivo killing by memory CD8 T cells. J Immunol 171, 27–31.
Liu, F., and Whitton, J. L. (2005) Cutting edge: re-evaluating the in vivo cytokine responses of CD8+ T cells during primary and secondary viral infections. J Immunol 174, 5936–40.
Kent, U. M. (1999) Purification of antibodies using ammonium sulfate fractionation or gel filtration. Methods Mol Biol 115, 11–8.
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Fang, M., Sigal, L. (2010). Studying NK Cell Responses to Ectromelia Virus Infections in Mice. In: Campbell, K. (eds) Natural Killer Cell Protocols. Methods in Molecular Biology, vol 612. Humana Press. https://doi.org/10.1007/978-1-60761-362-6_28
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DOI: https://doi.org/10.1007/978-1-60761-362-6_28
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