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Isolation and Characterization of Clostridium difficile Toxin-Specific Single-Domain Antibodies

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Single Domain Antibodies

Part of the book series: Methods in Molecular Biology ((MIMB,volume 911))

Abstract

Camelidae single-domain antibodies (VHHs) are a unique class of small binding proteins that are promising inhibitors of targets relevant to infection and immunity. With VHH selection from hyperimmunized phage display libraries now routine and the fact that VHHs possess long, extended complementarity-determining region (CDR3) loop structures that can access traditionally immunosilent epitopes, VHH-based inhibition of targets such as bacterial toxins are being explored. Toxin A and toxin B are high molecular weight exotoxins (308 kDa and 269 kDa, respectively) secreted by Clostridium difficile that are the causative agents of C. difficile-associated diseases in humans and in animals. Here, we provide protocols for the rapid generation of C. difficile toxin A- and toxin B-specific VHHs by llama immunization and recombinant antibody/phage display technology approaches and for further characterization of the VHHs with respect to toxin-binding affinity and specificity and the conformational nature of their epitopes.

This is National Research Council Canada Publication 50017.

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Notes

  1. 1.

    1  All phagemid and cloning vectors are freely available upon request.

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Acknowledgments

We thank Glenn Songer and Hien Trinh (Iowa State University) for providing us with purified C. difficile toxins and preparing the recombinant toxin fragments. The authors declare no financial conflict of interest.

Author information

Authors and Affiliations

  1. Institute for Biological Sciences, National Research Council Canada, Ottawa, ON, Canada

    Greg Hussack, Mehdi Arbabi-Ghahroudi, C. Roger MacKenzie & Jamshid Tanha

  2. Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada

    Greg Hussack

  3. School of Environmental Sciences, University of Guelph, Guelph, ON, Canada

    Mehdi Arbabi-Ghahroudi, C. Roger MacKenzie & Jamshid Tanha

  4. Department of Biology, Carleton University, Ottawa, ON, Canada

    Mehdi Arbabi-Ghahroudi

  5. Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ottawa, ON, Canada

    Jamshid Tanha

Authors
  1. Greg Hussack
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  2. Mehdi Arbabi-Ghahroudi
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  3. C. Roger MacKenzie
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  4. Jamshid Tanha
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Corresponding author

Correspondence to Jamshid Tanha .

Editor information

Editors and Affiliations

  1. , Molecular & Cellular Interactions, Vrije Universiteit Brussel, Pleinlaan 2, Brussels, 1050, Belgium

    Dirk Saerens

  2. , Molecular & Cellular Interactions, Vrije Universiteit Brussel, Pleinlaan 2, Brussels, 1050, Belgium

    Serge Muyldermans

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Hussack, G., Arbabi-Ghahroudi, M., MacKenzie, C.R., Tanha, J. (2012). Isolation and Characterization of Clostridium difficile Toxin-Specific Single-Domain Antibodies. In: Saerens, D., Muyldermans, S. (eds) Single Domain Antibodies. Methods in Molecular Biology, vol 911. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-968-6_14

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  • DOI: https://doi.org/10.1007/978-1-61779-968-6_14

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-967-9

  • Online ISBN: 978-1-61779-968-6

  • eBook Packages: Springer Protocols

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