Skip to main content

Phenotypic and Functional Analysis of Antigen-Specific T Cell Exhaustion

  • Protocol
  • First Online:
T-Cell Differentiation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1514))

Abstract

In some cancers and chronic infections exhausted T cells increase their expression of inhibitory receptors and demonstrate an impaired ability to produce cytokines and to proliferate. Immunological techniques such as MHC class I tetramer staining, intracellular cytokine staining, and CFSE dilution can be used to determine the memory status, inhibitory receptor expression, cytokine production, and proliferative capacity of antigen-specific CD8+ T cells. Here, we describe approaches to define the inhibitory receptor expression, cytokine production, and proliferative capacity of antigen-specific CD8+ T cells from HIV-infected and CMV-infected donors by using polychromatic flow cytometry.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 159.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Wherry EJ, Kurachi M (2015) Molecular and cellular insights into T cell exhaustion. Nat Rev Immunol 15(8):486–499. doi:10.1038/nri3862

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Kuchroo VK, Anderson AC, Petrovas C (2014) Coinhibitory receptors and CD8 T cell exhaustion in chronic infections. Curr Opin HIV AIDS 9(5):439–445. doi:10.1097/COH.0000000000000088

    Article  CAS  PubMed  Google Scholar 

  3. Yamamoto T, Price DA, Casazza JP et al (2011) Surface expression patterns of negative regulatory molecules identify determinants of virus-specific CD8+ T-cell exhaustion in HIV infection. Blood 117(18):4805–4815. doi:10.1182/blood-2010-11-317297

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Peretz Y, He Z, Shi Y et al (2012) CD160 and PD-1 co-expression on HIV-specific CD8 T cells defines a subset with advanced dysfunction. PLoS Pathog 8(8):e1002840. doi:10.1371/journal.ppat.1002840

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Freeman GJ, Long AJ, Iwai Y et al (2000) Engagement of the Pd-1 immunoinhibitory receptor by a novel B7 family member leads to negative regulation of lymphocyte activation. J Exp Med 192(7):1027–1034. doi:10.1084/jem.192.7.1027

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Latchman Y, Wood CR, Chernova T et al (2001) PD-L2 is a second ligand for PD-1 and inhibits T cell activation. Nat Immunol 2(3):261–268

    Article  CAS  PubMed  Google Scholar 

  7. Trautmann L, Janbazian L, Chomont N et al (2006) Upregulation of PD-1 expression on HIV-specific CD8+ T cells leads to reversible immune dysfunction. Nat Med 12(10):1198–1202. http://www.nature.com/nm/journal/v12/n10/suppinfo/nm1482_S1.html

  8. Day CL, Kaufmann DE, Kiepiela P et al (2006) PD-1 expression on HIV-specific T cells is associated with T-cell exhaustion and disease progression. Nature 443(7109):350–354. http://www.nature.com/nature/journal/v443/n7109/suppinfo/nature05115_S1.html

  9. Barber DL, Wherry EJ, Masopust D et al (2006) Restoring function in exhausted CD8 T cells during chronic viral infection. Nature 439(7077):682–687. http://www.nature.com/nature/journal/v439/n7077/suppinfo/nature04444_S1.html

  10. Velu V, Titanji K, Zhu B et al (2009) Enhancing SIV-specific immunity in vivo by PD-1 blockade. Nature 458(7235):206–210. http://www.nature.com/nature/journal/v458/n7235/suppinfo/nature07662_S1.html

  11. McDermott DF, Atkins MB (2013) PD-1 as a potential target in cancer therapy. Cancer Med 2(5):662–673. doi:10.1002/cam4.106

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Lamoreaux L, Roederer M, Koup R (2006) Intracellular cytokine optimization and standard operating procedure. Nat Protoc 1(3):1507–1516. doi:10.1038/nprot.2006.268

    Article  CAS  PubMed  Google Scholar 

  13. Chattopadhyay PK, Melenhorst JJ, Ladell K et al (2008) Techniques to improve the direct ex vivo detection of low frequency antigen-specific CD8+ T cells with peptide-major histocompatibility complex class I tetramers. Cytometry A 73A(11):1001–1009. doi:10.1002/cyto.a.20642

    Article  Google Scholar 

  14. Lyons AB, Blake SJ, Doherty KV (2001) Flow cytometric analysis of cell division by dilution of CFSE and related dyes, Current protocols in cytometry. Wiley, New York. doi:10.1002/0471142956.cy0911s64

    Google Scholar 

  15. Appay V, Rowland-Jones SL (2002) The assessment of antigen-specific CD8+ T cells through the combination of MHC class I tetramer and intracellular staining. J Immunol Methods 268(1):9–19. doi:10.1016/S0022-1759(02)00195-3

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Takuya Yamamoto Ph.D. .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2017 Springer Science+Business Media New York

About this protocol

Cite this protocol

Boswell, K.L., Yamamoto, T. (2017). Phenotypic and Functional Analysis of Antigen-Specific T Cell Exhaustion. In: Lugli, E. (eds) T-Cell Differentiation. Methods in Molecular Biology, vol 1514. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6548-9_6

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-6548-9_6

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6546-5

  • Online ISBN: 978-1-4939-6548-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics