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Absolute Quantification of Plasma Membrane Receptors Via Quantitative Flow Cytometry

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VEGF Signaling

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2475))

Abstract

Plasma membrane receptors are transmembrane proteins that initiate cellular response following the binding of specific ligands (e.g., growth factors, hormones, and cytokines). The abundance of plasma membrane receptors can be a diagnostic or prognostic biomarker in many human diseases. One of the best techniques for measuring plasma membrane receptors is quantitative flow cytometry (qFlow). qFlow employs fluorophore-conjugated antibodies against the receptors of interest and corresponding fluorophore-loaded calibration beads offers standardized and reproducible measurements of plasma membrane receptors. More importantly, qFlow can achieve absolute quantification of plasma membrane receptors when phycoerythrin (PE) is the fluorophore of choice. Here we describe a detailed qFlow protocol to obtain absolute receptor quantities on the basis of PE calibration. This protocol is foundational for many previous and ongoing studies in quantifying tyrosine kinase receptors and G-protein-coupled receptors with in vitro cell models and ex vivo cell samples.

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Acknowledgments

This work was supported by the American Heart Association (grant No. 16SDG26940002), National Science Foundation (grant No. 1653925), and National Institute of Child Health and Human Development (NIH grant No. R01HD096737). The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies.

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Authors and Affiliations

  1. Department of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO, USA

    Yingye Fang & P. I. Imoukhuede

  2. Center for Reproductive Health Sciences, Department of Obstetrics and Gynecology, Washington University School of Medicine, St. Louis, MO, USA

    Manasi Malik & Sarah K. England

  3. University of Washington, Department of Bioengineering, Seattle, WA, USA

    Yingye Fang

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  1. Yingye Fang
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  2. Manasi Malik
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  3. Sarah K. England
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  4. P. I. Imoukhuede
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Correspondence to P. I. Imoukhuede .

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Editors and Affiliations

  1. Radcliffe Department of Medicine, University of Oxford, Oxford, Oxfordshire, UK

    Lorna R. Fiedler

  2. Department of Comparative Biomedical Sciences, Royal Veterinary College, London, UK

    Caroline Pellet-Many

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Fang, Y., Malik, M., England, S.K., Imoukhuede, P.I. (2022). Absolute Quantification of Plasma Membrane Receptors Via Quantitative Flow Cytometry. In: Fiedler, L.R., Pellet-Many, C. (eds) VEGF Signaling. Methods in Molecular Biology, vol 2475. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2217-9_4

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  • DOI: https://doi.org/10.1007/978-1-0716-2217-9_4

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2216-2

  • Online ISBN: 978-1-0716-2217-9

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