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Detection of Cell Wall-Anchoring Machinery by Immunogold-Labeling Thin-Section Electron Microscopy

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The Bacterial Cell Wall

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2727))

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Abstract

Cell wall anchoring of surface proteins and pili in Gram-positive bacteria is mediated by sortase – a highly conserved transpeptidase enzyme. Early studies have demonstrated the membrane-associated nature of this enzyme in close proximity with its cognate substrates, using immunogold-labeling thin-section electron microscopy. Here, we provide a detail protocol of this methodology, including specimen preparation, ultrathin sectioning, and immunogold-labeling electron microscopic procedures, with an experimental model of sortase enzymes from Actinomyces oris. In principle, this protocol can be employed for any bacterial ultrathin-section samples to detect subcellular localization of proteins and organelles by immuno-electron microscopy.

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References

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Acknowledgments

This work was supported by the National Institute of Dental and Craniofacial Research of the National Institutes of Health under Award Number DE017382 (to H.T.-T.). The protocol was developed with assistance from the Department of Pathology at the University of Texas Health Science Center at Houston, Texas.

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Authors and Affiliations

  1. Division of Oral & Systemic Health Sciences, School of Dentistry, University of California, Los Angeles, CA, USA

    Chungyu Chang & Hung Ton-That

  2. Molecular Biology Institute, University of California, Los Angeles, CA, USA

    Hung Ton-That

  3. Department of Microbiology, Immunology & Molecular Genetics, University of California, Los Angeles, Los Angeles, CA, USA

    Hung Ton-That

Authors
  1. Chungyu Chang
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  2. Hung Ton-That
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Correspondence to Chungyu Chang .

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Editors and Affiliations

  1. Division of Oral & Systemic Health Sciences, School of Dentistry, University of California, Los Angeles, Los Angeles, CA, USA

    Hung Ton-That

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© 2024 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Chang, C., Ton-That, H. (2024). Detection of Cell Wall-Anchoring Machinery by Immunogold-Labeling Thin-Section Electron Microscopy. In: Ton-That, H. (eds) The Bacterial Cell Wall. Methods in Molecular Biology, vol 2727. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3491-2_12

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  • DOI: https://doi.org/10.1007/978-1-0716-3491-2_12

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3490-5

  • Online ISBN: 978-1-0716-3491-2

  • eBook Packages: Springer Protocols

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