Article | Published:

Parallel detection of antigen-specific T-cell responses by multidimensional encoding of MHC multimers

Nature Methods volume 6, pages 520526 (2009) | Download Citation

Subjects

Abstract

The use of fluorescently labeled major histocompatibility complex multimers has become an essential technique for analyzing disease- and therapy-induced T-cell immunity. Whereas classical major histocompatibility complex multimer analyses are well-suited for the detection of immune responses to a few epitopes, limitations on human-subject sample size preclude a comprehensive analysis of T-cell immunity. To address this issue, we developed a combinatorial encoding strategy that allows the parallel detection of a multitude of different T-cell populations in a single sample. Detection of T cells from peripheral blood by combinatorial encoding is as efficient as detection with conventionally labeled multimers but results in a substantially increased sensitivity and, most notably, allows comprehensive screens to be performed. We obtained proof of principle for the feasibility of large-scale screening of human material by analysis of human leukocyte antigen A3–restricted T-cell responses to known and potential melanoma-associated antigens in peripheral blood from individuals with melanoma.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    et al. A direct estimate of the human alphabeta T cell receptor diversity. Science 286, 958–961 (1999).

  2. 2.

    et al. Phenotypic analysis of antigen-specific T lymphocytes. Science 274, 94–96 (1996).

  3. 3.

    & MHC multimer technology: current status and future prospects. Curr. Opin. Immunol. 17, 428–433 (2005).

  4. 4.

    et al. Quantum dot semiconductor nanocrystals for immunophenotyping by polychromatic flow cytometry. Nat. Med. 12, 972–977 (2006).

  5. 5.

    , , & Selective expansion of cross-reactive CD8(+) memory T cells by viral variants. J. Exp. Med. 190, 1319–1328 (1999).

  6. 6.

    et al. Design and use of conditional MHC class I ligands. Nat. Med. 12, 246–251 (2006).

  7. 7.

    et al. Generation of peptide-MHC class I complexes through UV-mediated ligand exchange. Nat. Protoc. 1, 1120–1132 (2006).

  8. 8.

    et al. 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. USA 105, 3825–3830 (2008).

  9. 9.

    & The measurement of observer agreement for categorical data. Biometrics 33, 159–174 (1977).

  10. 10.

    et al. Memory CD8+ T cells vary in differentiation phenotype in different persistent virus infections. Nat. Med. 8, 379–385 (2002).

  11. 11.

    , , & Phenotype and function of human T lymphocyte subsets: consensus and issues. Cytometry A 73, 975–983 (2008).

  12. 12.

    et al. Identification of new melanoma epitopes on melanosomal proteins recognized by tumor infiltrating T lymphocytes restricted by HLA-A1, -A2, and -A3 alleles. J. Immunol. 161, 6985–6992 (1998).

  13. 13.

    et al. Identification of gp100-derived, melanoma-specific cytotoxic T-lymphocyte epitopes restricted by HLA-A3 supertype molecules by primary in vitro immunization with peptide-pulsed dendritic cells. Int. J. Cancer 78, 518–524 (1998).

  14. 14.

    et al. Shared epitopes for HLA-A3–restricted melanoma-reactive human CTL include a naturally processed epitope from Pmel-17/gp100. J. Immunol. 157, 5027–5033 (1996).

  15. 15.

    et al. Parasite stage-specific recognition of endogenous Toxoplasma gondii-derived CD8+ T cell epitopes. J. Infect. Dis. 198, 1625–1633 (2008).

  16. 16.

    , , , & The CD8 T-cell response against murine gammaherpesvirus 68 is directed toward a broad repertoire of epitopes from both early and late antigens. J. Virol. 82, 12205–12212 (2008).

  17. 17.

    et al. Discovery of CD8+ T cell epitopes in Chlamydia trachomatis infection through use of caged class I MHC tetramers. Proc. Natl. Acad. Sci. USA 105, 3831–3836 (2008).

  18. 18.

    , & HLA-A2–peptide complexes: refolding and crystallization of molecules expressed in Escherichia coli and complexed with single antigenic peptides. Proc. Natl. Acad. Sci. USA 89, 3429–3433 (1992).

  19. 19.

    et al. On the role of melanoma-specific CD8+ T-cell immunity in disease progression of advanced-stage melanoma patients. Clin. Cancer Res. 10, 4754–4760 (2004).

Download references

Acknowledgements

We thank B. Rodenko and H. Ovaa (Netherlands Cancer Institute) for the kind gift of J, W. van de Kasteele for help with cell culture, T. Wirenfeldt for statistical assistance, and A. Pfauth, F. van Diepen, M. van der Hoorn and G. de Roo for technical support with flow cytometry. This work was supported by the Danish Cancer Society grant DP06031 and the Carlsberg Foundation grant 2005-1-641 (to S.R.H.), Landsteiner Foundation of Blood Transfusion research grant 0522 and a Melanoma Research Alliance established investigator award (to T.N.S.) and Dutch Cancer Society grant UL 2007-3825 (to M.H.H. and T.N.M.S.).

Author information

Author notes

    • Sine Reker Hadrup
    •  & Arnold H Bakker

    Present addresses: Center for Cancer Immune Therapy, Department of Hematology, Herlev University Hospital, Herlev, Denmark (S.R.H.) and Division of Immunology and Pathogenesis, Department of Molecular and Cell Biology, University of California, Berkeley, USA (A.H.B.).

    • Sine Reker Hadrup
    •  & Arnold H Bakker

    These authors contributed equally to this work.

Affiliations

  1. Division of Immunology, The Netherlands Cancer Institute, Amsterdam, The Netherlands.

    • Sine Reker Hadrup
    • , Arnold H Bakker
    • , Chengyi J Shu
    • , Jerre van Veluw
    • , Emilie Castermans
    • , Christian Blank
    • , John B Haanen
    •  & Ton N Schumacher
  2. Center for Cancer Immune Therapy, Department of Hematology, Herlev University Hospital, Herlev, Denmark.

    • Rikke S Andersen
    •  & Per thor Straten
  3. Department of Hematology, Leiden University Medical Center, Leiden, The Netherlands.

    • Pleun Hombrink
    •  & Mirjam H Heemskerk

Authors

  1. Search for Sine Reker Hadrup in:

  2. Search for Arnold H Bakker in:

  3. Search for Chengyi J Shu in:

  4. Search for Rikke S Andersen in:

  5. Search for Jerre van Veluw in:

  6. Search for Pleun Hombrink in:

  7. Search for Emilie Castermans in:

  8. Search for Per thor Straten in:

  9. Search for Christian Blank in:

  10. Search for John B Haanen in:

  11. Search for Mirjam H Heemskerk in:

  12. Search for Ton N Schumacher in:

Contributions

S.R.H., A.H.B. and C.J.S. designed research, performed research, analyzed data and wrote the paper; R.S.A. performed research and analyzed data; J.v.V. performed research; P.H. and E.C. provided practical assistance; P.t.S., C.B. and J.B.H. contributed material from individuals with melanoma; M.H.H., designed research and analyzed data; T.N.S., designed research, analyzed data and wrote the paper.

Competing interests

The technology described in this manuscript is the subject of an EU patent application. Based on Netherlands Cancer Institute policy on management of intellectual property, S.R.H., A.H.B, C.J.S. and T.N.S would be entitled to a portion of received royalty income.

Corresponding author

Correspondence to Ton N Schumacher.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–4 and Supplementary Tables 1–4

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nmeth.1345

Further reading

  • Glutaminyl cyclase is an enzymatic modifier of the CD47- SIRPα axis and a target for cancer immunotherapy

    • Meike E. W. Logtenberg
    • , J. H. Marco Jansen
    • , Matthijs Raaben
    • , Mireille Toebes
    • , Katka Franke
    • , Arianne M. Brandsma
    • , Hanke L. Matlung
    • , Astrid Fauster
    • , Raquel Gomez-Eerland
    • , Noor A. M. Bakker
    • , Simone van der Schot
    • , Koen A. Marijt
    • , Martijn Verdoes
    • , John B. A. G. Haanen
    • , Joost H. van den Berg
    • , Jacques Neefjes
    • , Timo K. van den Berg
    • , Thijn R. Brummelkamp
    • , Jeanette H. W. Leusen
    • , Ferenc A. Scheeren
    •  & Ton N. Schumacher

    Nature Medicine (2019)

  • Induction of neoantigen-reactive T cells from healthy donors

    • Muhammad Ali
    • , Zsofia Foldvari
    • , Eirini Giannakopoulou
    • , Maxi-Lu Böschen
    • , Erlend Strønen
    • , Weiwen Yang
    • , Mireille Toebes
    • , Benjamin Schubert
    • , Oliver Kohlbacher
    • , Ton N. Schumacher
    •  & Johanna Olweus

    Nature Protocols (2019)

  • Immune biomarkers for predicting response to adoptive cell transfer as cancer treatment

    • Ianthe A. E. M. van Belzen
    •  & Can Kesmir

    Immunogenetics (2019)

  • Actively personalized vaccination trial for newly diagnosed glioblastoma

    • Norbert Hilf
    • , Sabrina Kuttruff-Coqui
    • , Katrin Frenzel
    • , Valesca Bukur
    • , Stefan Stevanović
    • , Cécile Gouttefangeas
    • , Michael Platten
    • , Ghazaleh Tabatabai
    • , Valerie Dutoit
    • , Sjoerd H. van der Burg
    • , Per thor Straten
    • , Francisco Martínez-Ricarte
    • , Berta Ponsati
    • , Hideho Okada
    • , Ulrik Lassen
    • , Arie Admon
    • , Christian H. Ottensmeier
    • , Alexander Ulges
    • , Sebastian Kreiter
    • , Andreas von Deimling
    • , Marco Skardelly
    • , Denis Migliorini
    • , Judith R. Kroep
    • , Manja Idorn
    • , Jordi Rodon
    • , Jordi Piró
    • , Hans S. Poulsen
    • , Bracha Shraibman
    • , Katy McCann
    • , Regina Mendrzyk
    • , Martin Löwer
    • , Monika Stieglbauer
    • , Cedrik M. Britten
    • , David Capper
    • , Marij J. P. Welters
    • , Juan Sahuquillo
    • , Katharina Kiesel
    • , Evelyna Derhovanessian
    • , Elisa Rusch
    • , Lukas Bunse
    • , Colette Song
    • , Sandra Heesch
    • , Claudia Wagner
    • , Alexandra Kemmer-Brück
    • , Jörg Ludwig
    • , John C. Castle
    • , Oliver Schoor
    • , Arbel D. Tadmor
    • , Edward Green
    • , Jens Fritsche
    • , Miriam Meyer
    • , Nina Pawlowski
    • , Sonja Dorner
    • , Franziska Hoffgaard
    • , Bernhard Rössler
    • , Dominik Maurer
    • , Toni Weinschenk
    • , Carsten Reinhardt
    • , Christoph Huber
    • , Hans-Georg Rammensee
    • , Harpreet Singh-Jasuja
    • , Ugur Sahin
    • , Pierre-Yves Dietrich
    •  & Wolfgang Wick

    Nature (2019)

  • Combinatorial detection of autoreactive CD8+ T cells with HLA-A2 multimers: a multi-centre study by the Immunology of Diabetes Society T Cell Workshop

    • Eddie A. James
    • , Joana R. F. Abreu
    • , John W. McGinty
    • , Jared M. Odegard
    • , Yvonne E. Fillié
    • , Claire N. Hocter
    • , Slobodan Culina
    • , Kristin Ladell
    • , David A. Price
    • , Aimon Alkanani
    • , Marynette Rihanek
    • , Lisa Fitzgerald-Miller
    • , Ania Skowera
    • , Cate Speake
    • , Peter Gottlieb
    • , Howard W. Davidson
    • , F. Susan Wong
    • , Bart Roep
    •  & Roberto Mallone

    Diabetologia (2018)