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Measurement of Salmonella enterica Internalization and Vacuole Lysis in Epithelial Cells

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Phagocytosis and Phagosomes

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2692))

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Abstract

Establishment of an intracellular niche within mammalian cells is key to the pathogenesis of the gastrointestinal bacterium, Salmonella enterica serovar Typhimurium (S. Typhimurium). Here we will describe how to study the internalization of S. Typhimurium into human epithelial cells using the gentamicin protection assay. The assay takes advantage of the relatively poor penetration of gentamicin into mammalian cells; internalized bacteria are effectively protected from its antibacterial actions. A second assay, the chloroquine (CHQ) resistance assay, can be used to determine the proportion of internalized bacteria that have lysed or damaged their Salmonella-containing vacuole and are therefore residing within the cytosol. Its application to the quantification of cytosolic S. Typhimurium in epithelial cells will also be presented. Together, these protocols provide an inexpensive, rapid, and sensitive quantitative measure of bacterial internalization and vacuole lysis by S. Typhimurium.

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Acknowledgments

This work was supported by start-up funds from the Paul G. Allen School for Global Animal Health and the Stanley L. Adler Research Fund (LAK). TRP is supported by an NIH/NIAID Training Grant 2T32AI007025. JAK is the recipient of a Poncin Scholarship.

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Authors and Affiliations

  1. Paul G. Allen School for Global Health, College of Veterinary Medicine, Washington State University, Pullman, WA, USA

    Jessica A. Klein, TuShun R. Powers & Leigh A. Knodler

  2. Department of Microbiology and Molecular Genetics, University of Vermont, Burlington, USA

    Leigh A. Knodler

Authors
  1. Jessica A. Klein
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  2. TuShun R. Powers
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  3. Leigh A. Knodler
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Correspondence to Leigh A. Knodler .

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Editors and Affiliations

  1. Department of Chemistry and Biology, Toronto Metropolitan University, Toronto, ON, Canada

    Roberto J. Botelho

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Klein, J.A., Powers, T.R., Knodler, L.A. (2023). Measurement of Salmonella enterica Internalization and Vacuole Lysis in Epithelial Cells. In: Botelho, R.J. (eds) Phagocytosis and Phagosomes. Methods in Molecular Biology, vol 2692. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3338-0_14

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  • DOI: https://doi.org/10.1007/978-1-0716-3338-0_14

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3337-3

  • Online ISBN: 978-1-0716-3338-0

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