Fluorescent erythrocyte ghosts as standards for quantitative flow cytometry

Authors

  • S. K. Doberstein,

    1. Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland
    Current affiliation:
    1. Department of Molecular and Cell Biology, Division of Neuroscience, 519 LSA, University of California, Berkeley, Berkeley, CA 94720. Address reprint requests there
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  • G. Wiegand,

    1. Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland
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  • L. M. Machesky,

    1. Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland
    Current affiliation:
    1. Medical Research Council Laboratory of Molecular Biology, Cambridge, England
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  • T. D. Pollard

    1. Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland
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Abstract

We report here a quick and inexpensive method for preparing standards of known fluorochrome content for calibration and quantitation of flow cytometry fluorescence signals. Erythrocyte ghosts prepared by hypotonic lysis are filled with solutions containing fluorescently labeled dextran. Standards prepared by this technique have a narrow range of fluorescence and a linear response of fluorescence to fluorochrome content up to 2 × 106 fluorochrome molecules/cell. The volume of ghost standard particles is roughly 70 femtoliters (fl)/cell. The fluorescence of ghost standards is nearly identical to that of commercially available microbead standards of similar fluorochrome content. Ghost standards have stable fluorescence for at least 3 weeks at 4°C. These standards can be made with any fluorochrome or combination of fluorochromes over a wide concentration range. © 1995 Wiley-Liss, Inc.

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