Summary
Recombinant major histocompatibility complex (MHC) class I molecules complexed with pathogen-specific or other disease-associated antigens have become essential reagents for the analysis of adaptive T-cell responses. However, conventional techniques for the production of recombinant peptide-MHC (pMHC) complexes are highly involved and thereby limit the use of pMHC complexes in terms of antigen diversity. To make pMHC-based techniques suitable for high-throughput analyses we developed an MHC peptide exchange technology based on the use of conditional MHC ligands. This technology enables the parallel production of thousands of different pMHC complexes within hours, allowing the development of high-throughput MHC-based assay systems to identify MHC ligands and cytotoxic T-cell responses. These high-throughput assays should prove valuable for the screening of entire disease-associated proteomes, including pathogen-encoded proteomes, tumor-associated antigens, and autoimmune antigens.
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
Arstila, T. P., Casrouge, A., Baron, V., Even, J., Kanellopoulos, J., and Kourilsky, P. (1999) A direct estimate of the human alphabeta T cell receptor diversity. Science 286, 958–961.
Altman, J. D., Moss, P. A., Goulder, P. J., Barouch, D. H., McHeyzer-Williams, M. G., Bell, J. I., McMichael, A. J., and Davis, M. M. (1996) Phenotypic analysis of antigen-specific T lymphocytes. Science 274, 94–96.
Toebes, M., Coccoris, M., Bins, A. D., Rodenko, B., Gomez, R., Nieuwkoop, N. J., van de Kasteele, W., Rimmelzwaan, G., Haanen, J. B., and Schumacher, T. N. (2006) Design and use of conditional MHC class I ligands. Nat. Med. 12, 246–251.
Checovich, W. J., Bolger, R. E., and Burke, T. (1995) Fluorescence polarization–a new tool for cell and molecular biology. Nature 375, 254–256.
Cobbold, M., Khan, N., Pourgheysari, B., Tauro, S., McDonald, D., Osman, H., Assenmacher, M., Billingham, L., Steward, C., Crawley, C., Olavarria, E., Goldman, J., Chakraverty, R., Mahendra, P., Craddock, C., and Moss, P. A. (2005) Adoptive transfer of cytomegalovirus-specific CTL to stem cell transplant patients after selection by HLA-peptide tetramers.J. Exp. Med. 202, 379–386.
Dudley, M. E., Wunderlich, J. R., Yang, J. C., Sherry, R. M., Topalian, S. L., Restifo, N. P., Royal, R. E., Kammula, U., White, D. E., Mavroukakis, S. A., Rogers, L. J., Gracia, G. J., Jones, S. A., Mangiameli, D. P., Pelletier, M. M., Gea-Banacloche, J., Robinson, M. R., Berman, D. M., Filie, A. C., Abati, A., and Rosenberg, S. A. (2005) Adoptive cell transfer therapy following non-myeloablative but lymphodepleting chemotherapy for the treatment of patients with refractory metastatic melanoma. J. Clin. Oncol. 23, 2346–2357.
Bakker, A. H. and Schumacher, T. N. (2005) MHC multimer technology: current status and future prospects. Curr. Opin. Immunol. 17, 428–433.
Grotenbreg, G. M., Nicholson, M. J., Fowler, K. D., Wilbuer, K., Octavio, L., Yang, M., Chakraborty, A. K., Ploegh, H. L., and Wucherpfennig, K. W. (2007) Empty class II major histocompatibility complex created by peptide photolysis establishes the role of DM in peptide association. J. Biol. Chem. 282, 21425–21436.
Bakker, A. H., Hoppes, R., Linnemann, C., Toebes, M., Rodenko, B., Berkers, C. R., Hadrup, S. R., van Esch, W. J., Heemskerk, M. H., Ovaa, H., and Schumacher, T. N. (2008) Conditional MHC class I ligands and peptide exchange technology for the human MHC gene products HLA-A1, -A3, -A11 and -B7. Proc. Natl. Acad. Sci. U. S. A. 105, 3825–3830.
Rodenko, B., Toebes, M., Hadrup, S. R., van Esch, W. J., Molenaar, A. M., Schumacher, T. N., and Ovaa, H. (2006) Generation of peptide-MHC class I complexes through UV-mediated ligand exchange. Nat. Protoc. 1, 1120–1132.
Wellings, D. A. and Atherton, E. (1997) Standard Fmoc protocols. Meth. Enzymol. 289, 44–67.
Garboczi, D. N., Hung, D. T., and Wiley, D. C. (1992) HLA-A2-peptide complexes: refolding and crystallization of molecules expressed in Escherichia coli and complexed with single antigenic peptides. Proc. Natl. Acad. Sci. U. S. A. 89, 3429–3433.
Coulie, P. G., Brichard, V., van Pel, A., Wolfel, T., Schneider, J., Traversari, C., Mattei, S., De Plaen, E., Lurquin, C., Szikora, J. P., Renauld, J. C., and Boon, T. (1994) A new gene coding for a differentiation antigen recognized by autologous cytolytic T lymphocytes on HLA-A2 melanomas. J. Exp. Med. 180, 35–42.
Andersen, M. H., Pedersen, L. O., Becker, J. C., and thor Straten, P. (2001) Identification of a cytotoxic T lymphocyte response to the apoptosis inhibitor protein surviving in cancer patients. Cancer Res. 61, 869–872.
Bruchez, M. Jr., Moronne, M., Gin, P., Weiss, S., and Alivisatos, A. P. (1998) Semiconductor nanocrystals as fluorescent biological labels. Science 281, 2013–2016.
Chattopadhyay, P. K., Price, D. A., Harper, T. F., Betts, M. R., Yu, J., Gostick, E., Perfetto, S. P., Goepfert, P., Koup, R. A., De Rosa, S. C., Bruchez, M. P., and Roederer, M. (2006) Quantum dot semiconductor nanocrystals for immunophenotyping by polychromatic flow cytometry. Nat. Med. 12, 972–977.
Soen, Y., Chen, D. S., Kraft, D. L., Davis, M. M., and Brown, P. O. (2003) Detection and characterization of cellular immune responses using peptide-MHC microarrays. PLoS Biol. 1, 429–438.
Chen, D. S., Soen, Y., Stuge, T. B., Lee, P. P., Weber, J. S., Brown, P. O., and Davis, M. M. (2005) Marked differences in human melanoma antigen-specific T cell responsiveness after vaccination using a functional microarray. PLoS Med. 2, 1018–1030.
Stone, J. D., Demkowicz, W. E. Jr., and Stern, L. J. (2005) HLA-restricted epitope identification and detection of functional T cell responses by using MHC-peptide and costimulatory microarrays. Proc. Natl. Acad. Sci. U. S. A. 102, 3744–3749.
Deviren, G., Gupta, K., Paulaitis, M. E., and Schneck, J. P. (2007) Detection of antigen-specific T cells on p/MHC microarrays. J. Mol. Recognit. 20, 32–38.
Guillaume, P., Legler, D. F., Boucheron, N., Doucey, M. A., Cerottini, J. C., and Luescher, I. F. (2003) Soluble major histocompatibility complex-peptide octamers with impaired CD8 binding selectively induce Fas-dependent apoptosis. J. Biol. Chem. 278, 4500–4509.
Ramachandiran, V., Grigoriev, V., Lan, L., Ravkov, E., Mertens, S. A., and Altman, J. D. (2007) A robust method for production of MHC tetramers with small molecule fluorophores. J. Immunol. Meth. 319, 13–20.
Grotenbreg, G. M., Roan, N. R., Guillen, E., Meijers, R., Wang, J. H., Bell, G. W., Starnbach, M. N., and Ploegh H. L. (2008) Discovery of CD8+ T cell epitopes in Chlamydia trachomatis infection through use of caged class I MHC tetramers. Proc. Natl. Acad. Sci. U. S. A. 105, 3831–3836.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2009 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Hadrup, S.R. et al. (2009). High-Throughput T-Cell Epitope Discovery Through MHC Peptide Exchange. In: Schutkowski, M., Reineke, U. (eds) Epitope Mapping Protocols. Methods in Molecular Biology™, vol 524. Humana Press. https://doi.org/10.1007/978-1-59745-450-6_28
Download citation
DOI: https://doi.org/10.1007/978-1-59745-450-6_28
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-934115-17-6
Online ISBN: 978-1-59745-450-6
eBook Packages: Springer Protocols