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High-Dimensional Flow Cytometry Analysis of Regulatory Receptors on Human T Cells, NK Cells, and NKT Cells

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Translational Bioinformatics for Therapeutic Development

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2194))

Abstract

The field of flow cytometry has witnessed rapid technological advancements in the last few decades. While the founding principles of fluorescent detection on cells (or particles) within a uniform fluid stream remains largely unchanged, the availability more sensitive cytometers with the ability to multiplex more and more florescent signals has resulted in very complex high-order assays. This results in the co-use of fluorophores with increased levels of emission overlap and/or spillover spreading than in years past and thus requires careful and well thought out planning for flow cytometry assay development. As an example, we present the development of a large 18-color (20 parameter) flow cytometry assay designed to take an in depth analysis of effector lymphocyte phenotypes, with careful attention to assay controls and panel design.

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Acknowledgments

This work was supported by the Moffitt Cancer Center—Innovative Core Projects (Project number 16060201), NCI–NIH (1 R01 CA148995-01; P30CA076292; P50CA168536), the V Foundation, the Dr. Miriam and Sheldon G. Adelson Medical Research Foundation, and the Chris Sullivan Foundation.

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Authors and Affiliations

  1. Department of Immunology, H. Lee Moffitt Cancer Center & Research Institute, Tampa, FL, USA

    Ryosuke Nakagawa, Nicole Restrepo, James J. Mulé & Adam W. Mailloux

  2. Department of Malignant Hematology, H. Lee Moffitt Cancer Center & Research Institute, Tampa, FL, USA

    Jason Brayer

  3. Department of Cutaneous Oncology, H. Lee Moffitt Cancer Center, Tampa, FL, USA

    James J. Mulé

Authors
  1. Ryosuke Nakagawa
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  2. Jason Brayer
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  3. Nicole Restrepo
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  4. James J. Mulé
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  5. Adam W. Mailloux
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Corresponding author

Correspondence to Adam W. Mailloux .

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Editors and Affiliations

  1. Department of Cutaneous Oncology, H. Lee Moffitt Cancer Center & Research Institute, Tampa, FL, USA

    Joseph Markowitz

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Nakagawa, R., Brayer, J., Restrepo, N., Mulé, J.J., Mailloux, A.W. (2021). High-Dimensional Flow Cytometry Analysis of Regulatory Receptors on Human T Cells, NK Cells, and NKT Cells. In: Markowitz, J. (eds) Translational Bioinformatics for Therapeutic Development. Methods in Molecular Biology, vol 2194. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0849-4_14

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  • DOI: https://doi.org/10.1007/978-1-0716-0849-4_14

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0848-7

  • Online ISBN: 978-1-0716-0849-4

  • eBook Packages: Springer Protocols

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