Skip to main content
SpringerLink
Log in

Intracellular Staining for Cytokines and Transcription Factors

  • Protocol
  • First Online:
T-Helper Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1193))

Abstract

Within the past years immune cells in general and T cells in particular have been categorized into a vast variety of subsets with different functional properties. One of the key technologies fueling this emerging complexity is intracellular staining for effector cytokines and/or lineage-defining transcription factors. Here we discuss the critical steps for performing successful multicolor immunophenotyping of mouse T cells in combination with analysis of intracellular molecules after ex vivo isolation.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
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. Perfetto SP, Chattopadhyay PK, Roederer M (2004) Seventeen-colour flow cytometry: unravelling the immune system. Nat Rev Immunol 4:648–655

    Article  CAS  PubMed  Google Scholar 

  2. Mahnke YD, Roederer M (2007) Optimizing a multicolor immunophenotyping assay. Clin Lab Med 27:469–485, v

    Article  PubMed Central  PubMed  Google Scholar 

  3. Roederer M (2001) Spectral compensation for flow cytometry: visualization artifacts, limitations, and caveats. Cytometry 45:194–205

    Article  CAS  PubMed  Google Scholar 

  4. Sander B, Andersson J, Andersson U (1991) Assessment of cytokines by immunofluorescence and the paraformaldehyde-saponin procedure. Immunol Rev 119:65–93

    Article  CAS  PubMed  Google Scholar 

  5. Jung T, Schauer U, Heusser C, Neumann C, Rieger C (1993) Detection of intracellular cytokines by flow cytometry. J Immunol Methods 159:197–207

    Article  CAS  PubMed  Google Scholar 

  6. Bluestone JA, Mackay CR, O’Shea JJ, Stockinger B (2009) The functional plasticity of T cell subsets. Nat Rev Immunol 9:811–816

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Zhu J, Yamane H, Paul WE (2010) Differentiation of effector CD4 T cell populations *. Annu Rev Immunol 28:445–489

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  8. Hirota K et al (2011) Fate mapping of IL-17-producing T cells in inflammatory responses. Nat Immunol 12:255–263

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  9. Aghaeepour N et al (2013) Critical assessment of automated flow cytometry data analysis techniques. Nat Meth 10:228–238

    Article  CAS  Google Scholar 

  10. Perfetto SP et al. (2010) Amine-reactive dyes for dead cell discrimination in fixed samples. Curr Protoc Cytom Chapter 9, Unit 9.34

    Google Scholar 

  11. Brosterhus H et al (1999) Enrichment and detection of live antigen-specific CD4(+) and CD8(+) T cells based on cytokine secretion. Eur J Immunol 29:4053–4059

    Article  CAS  PubMed  Google Scholar 

  12. Manz R, Assenmacher M, Pflüger E, Miltenyi S, Radbruch A (1995) Analysis and sorting of live cells according to secreted molecules, relocated to a cell-surface affinity matrix. Proc Natl Acad Sci U S A 92:1921–1925

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  13. Codarri L et al (2011) RORγt drives production of the cytokine GM-CSF in helper T cells, which is essential for the effector phase of autoimmune neuroinflammation. Nat Immunol 12:560–567

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Institute of Experimental Immunology, University of Zürich, Winterthurerstrasse 190, 8057, Zürich, Switzerland

    Florian Mair & Vinko Tosevski

Authors
  1. Florian Mair
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Vinko Tosevski
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Florian Mair .

Editor information

Editors and Affiliations

  1. Institute for Molecular Medicine, University Medical Center of the Johannes Gutenberg University of Mainz, Mainz, Germany

    Ari Waisman

  2. University of Zurich Institute of Experimental Immunology, Zurich, Germany

    Burkhard Becher

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media New York

About this protocol

Cite this protocol

Mair, F., Tosevski, V. (2014). Intracellular Staining for Cytokines and Transcription Factors. In: Waisman, A., Becher, B. (eds) T-Helper Cells. Methods in Molecular Biology, vol 1193. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1212-4_5

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-1212-4_5

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1211-7

  • Online ISBN: 978-1-4939-1212-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation