Abstract
The advent of soluble MHC multimer technology has allowed for the flow-cytometric direct identification of specific-MHC restricted antigen-specific T cells in mixed cell populations and also enabled the direct phenotyping and cloning of these cells at the same time. To date, MHC multimers have been used in characterizing the adaptive T cell repertoire under infectious, cancerous, and autoimmune states and has increased our understanding of the dynamics of T-cell immunity. Recombinant MHC multimers have been produced where MHC-binding peptide antigens are either covalently or noncovalently bound to the MHC, with the latter having the advantage of the ability to use a single recombinant MHC to investigate multiple MHC-binding peptides and their interacting T cells. In this method we describe how to generate recombinant non-covalently bound peptide MHC-multimers in insect cells. MHC multimers are generated as tetravalent complexes using a streptavidin scaffold.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Day, C.L., Seth, N.P., Lucas, M., et al. (2003) Ex vivo analysis of human memory CD4 T cells specific for hepatitis C virus using MHC class II tetramers. J.Clin.Invest 112, 831–842.
Novak, E.J., Liu, A.W., Nepom, G.T., and Kwok, W.W. (1999) MHC class II tetramers identify peptide-specific human CD4(+) T cells proliferating in response to influenza A antigen. J Clin Invest 104, R63–R67.
Danke, N.A. and Kwok, W.W. (2003) HLA class II-restricted CD4+ T cell responses directed against influenza viral antigens postinfluenza vaccination. J.Immunol. 171, 3163–3169.
Bischof, F., Hofmann, M., Schumacher, T.N., et al. (2004) Analysis of autoreactive CD4 T cells in experimental autoimmune encephalomyelitis after primary and secondary challenge using MHC class II tetramers. J.Immunol. 172, 2878–2884.
Novak, E.J., Liu, A.W., Gebe, J.A., et al. (2001) Tetramer-guided epitope mapping: rapid identification and characterization of immunodominant CD4+ T cell epitopes from complex antigens. J.Immunol. 166, 6665–6670.
Reijonen, H., Novak, E.J., Kochik, S., et al. (2002) Detection of GAD65 specific T-cells by MHC class II multimers in type 1 diabetes patients and at-risk subjects. Diabetes 51, 1375–1382.
Altman, J.D., Moss, P.A., Goulder, P.J., et al. (1996) Phenotypic analysis of antigen-specific T lymphocytes. Science 274, 94–96.
Crawford, F., Kozono, H., White, J., Marrack, P., and Kappler, J. (1998) Detection of antigen-specific T cells with multivalent soluble class II MHC covalent peptide complexes. Immunity 8, 675–682.
Appel, H., Gauthier, L., Pyrdol, J., and Wucherpfennig, K.W. (2000) Kinetics of T-cell receptor binding by bivalent HLA-DR.Peptide complexes that activate antigen-specific human T-cells. J.Biol.Chem. 275, 312–321.
Gutgemann, I., Fahrer, A.M., Altman, J.D., Davis, M.M., and Chien, Y.H. (1998) Induction of rapid T cell activation and tolerance by systemic presentation of an orally administered antigen. Immunity. 8, 667–673.
Mallet-Designe, V.I., Stratmann, T., Homann, D., Carbone, F., Oldstone, M.B., and Teyton, L. (2003) Detection of low-avidity CD4+ T cells using recombinant artificial APC: following the antiovalbumin immune response. J.Immunol. 170, 123–131.
Cochran, J.R., Cameron, T.O., and Stern, L.J. (2000) The relationship of MHCpeptide binding and T cell activation probed using chemically defined MHC class II oligomers. Immunity 12, 241–250.
Malherbe, L., Filippi, C., Julia, V., et al. (2000) Selective activation and expansion of high-affinity CD4+ T cells in resistant mice upon infection with Leishmania major. Immunity. 13, 771–782.
Arnold, P.Y., Vignali, K.M., Miller, T.B., et al. (2002) Reliable generation and use of MHC class II:gamma2aFc multimers for the identification of antigen-specific CD4(+) T cells. J.Immunol.Methods 271, 137–151.
Jang, M.H., Seth, N.P., and Wucherpfennig, K.W. (2003) Ex vivo analysis of thymic CD4 T cells in nonobese diabetic mice with tetramers generated from IA(g7)/class II-associated invariant chain peptide precursors. J.Immunol. 171, 4175–4186.
Buckner, J.H., Van, L.M., Kwok, W.W., and Tsarknaridis, L. (2002) Identification of type II collagen peptide 261–273-specific T cell clones in a patient with relapsing polychondritis. Arthritis Rheum. 46, 238–244.
Kotzin, B.L., Falta, M.T., Crawford, F., et al. (2000) Use of soluble peptide-DR4 tetramers to detect synovial T cells specific for cartilage antigens in patients with rheumatoid arthritis. Proc.Natl.Acad.Sci.USA 97, 291–296.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2007 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Gebe, J.A., Kwok, W.W. (2007). Tracking Antigen Specific CD4+ T-Cells With Soluble MHC Molecules. In: Cope, A.P. (eds) Arthritis Research. Methods in Molecular Medicine, vol 136. Humana Press. https://doi.org/10.1007/978-1-59745-402-5_4
Download citation
DOI: https://doi.org/10.1007/978-1-59745-402-5_4
Publisher Name: Humana Press
Print ISBN: 978-1-58829-918-5
Online ISBN: 978-1-59745-402-5
eBook Packages: Springer Protocols