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Guidelines for Gating Flow Cytometry Data for Immunological Assays

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Immunophenotyping

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2032))

Abstract

“Gating” refers to the selection of successive subpopulations of cells for analysis in flow cytometry. It is usually performed manually, based on expert knowledge of cell characteristics. However, there can be considerable disagreement in how gates should be applied, even between individuals experienced in the field. While clinical software often automates gating, and some guidelines do exist (especially for clinical assays), there are no comprehensive guidelines across the various types of immunological assays performed using flow cytometry. Here we attempt to provide such guidelines, focused on the most general and pervasive types of gates, why they are important, and what recommendations can be made regarding their use. We do so through the display of example data, collected by academic, government, and industry representatives. These guidelines should be of value to both novice and experienced flow cytometrists analyzing a wide variety of immunological assays.

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Acknowledgments

The authors thank the EQAPOL consortium and Jennifer Enzor for providing data and analysis examples, and the FOCIS Immunophenotyping Course participants for helpful suggestions and vetting of these guidelines.

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Authors and Affiliations

  1. Duke Immune Profiling Core, Duke University Medical Center, Durham, NC, USA

    Janet Staats

  2. Department for Cellular Analysis, Biolegend, San Diego, CA, USA

    Anagha Divekar

  3. Frederick, MD, USA

    J. Philip McCoy Jr.

  4. Institute for Immunity, Transplantation, and Infection, Stanford University School of Medicine, Stanford, CA, USA

    Holden T. Maecker

Authors
  1. Janet Staats
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  2. Anagha Divekar
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  3. J. Philip McCoy Jr.
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  4. Holden T. Maecker
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Corresponding author

Correspondence to Holden T. Maecker .

Editor information

Editors and Affiliations

  1. Frederick, MD, USA

    J. Philip McCoy, Jr

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Staats, J., Divekar, A., McCoy, J.P., Maecker, H.T. (2019). Guidelines for Gating Flow Cytometry Data for Immunological Assays. In: McCoy, Jr, J. (eds) Immunophenotyping. Methods in Molecular Biology, vol 2032. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9650-6_5

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  • DOI: https://doi.org/10.1007/978-1-4939-9650-6_5

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9649-0

  • Online ISBN: 978-1-4939-9650-6

  • eBook Packages: Springer Protocols

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