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Multiparameter Intracellular Cytokine Staining

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Flow Cytometry Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1678))

Abstract

Intracellular cytokine staining is a popular method for visualizing cellular responses, most often T-cell responses to antigenic or mitogenic stimulation. It can be coupled with staining for other functional markers, such as upregulation of CD107 or CD154, as well as phenotypic markers that define specific cellular subsets, e.g., effector and memory T-cell compartments, NK cells, or monocytes. Recent advances in multicolor flow cytometry instrumentation and software have allowed the routine combination of 12 or more markers, creating some technical and analytical challenges along the way, and exposing a need for standardization in the field. Here, we will review best practices for antibody panel design and procedural variables for multicolor intracellular cytokine staining, and present an optimized protocol with variations designed for use with specific markers and sample types.

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Acknowledgments

Details of this protocol were optimized by Laurel Nomura and Maria Suni (BD Biosciences).

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Authors and Affiliations

  1. Human Immune Monitoring Center, Institute for Immunity, Transplantation, and Infection, Stanford University, Fairchild Science Building, 299 Campus Drive, Stanford, CA, 94305-5124, USA

    Patricia Lovelace & Holden T. Maecker

Authors
  1. Patricia Lovelace
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  2. Holden T. Maecker
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Corresponding author

Correspondence to Holden T. Maecker .

Editor information

Editors and Affiliations

  1. Flow Cytometry Section, Research Technologies Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA

    Teresa S. Hawley

  2. Department of Anatomy and Regenerative Biology, School of Medicine and Health Sciences, George Washington University, Washington, District of Columbia, USA

    Robert G. Hawley

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Lovelace, P., Maecker, H.T. (2018). Multiparameter Intracellular Cytokine Staining. In: Hawley, T., Hawley, R. (eds) Flow Cytometry Protocols. Methods in Molecular Biology, vol 1678. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7346-0_9

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  • DOI: https://doi.org/10.1007/978-1-4939-7346-0_9

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7344-6

  • Online ISBN: 978-1-4939-7346-0

  • eBook Packages: Springer Protocols

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