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Quorum Sensing pp 325-352 | Cite as

Generation of High-Sensitivity Monoclonal Antibodies Specific for Homoserine Lactones

  • Soumya Palliyil
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1673)

Abstract

A number of bacteria use a class of chemical compounds called acyl-homoserine lactones (AHLs) as quorum sensing (QS) signals to coordinate their behavior at the population level, including pathogens like Pseudomonas aeruginosa. Blocking QS using antibodies is an attractive strategy for infection control as this process takes a central role in P. aeruginosa infections. Here the methods involved in the generation of high sensitivity anti-QS monoclonal antibodies from an immunized sheep phage display antibody library are described. A panel of AHL compounds conjugated to carrier proteins are used for sheep immunization and a phage display antibody library is constructed using the immune repertoire of sheep as a source of antibody genes. High sensitivity single chain antibody fragments (scFv) are isolated from the library using “smart selection strategies” and reformatted into single chain antibodies (scAbs). The resultant monoclonal antibodies: (1) recognize HSL compounds at low nanomolar concentrations; (2) have the potential to reduce virulence gene expression in P. aeruginosa; and (3) offer protection in a nematode model of infection.

Key words

Pseudomonas aeruginosa Quorum sensing Acyl-homoserine lactones Monoclonal antibodies Anti-infective antibodies 

References

  1. 1.
    de Kievit TR, Iglewski BH (2000) Bacterial quorum sensing in pathogenic relationships. Infect Immun 68:4839–4849CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Palliyil S, Broadbent ID (2009) Novel immunotherapeutic approaches to the treatment of infections caused by Gram-negative bacteria. Curr Opin Pharmacol 9:566–570CrossRefPubMedGoogle Scholar
  3. 3.
    Passador L, Cook JM, Gambello MJ, Rust L, Iglewski BH (1993) Expression of Pseudomonas aeruginosa virulence genes requires cell-to-cell communication. Science 260:1127–1130CrossRefPubMedGoogle Scholar
  4. 4.
    Kaufmann GF, Park J, Mee JM, Ulevitch RJ, Janda KD (2008) The quorum quenching antibody RS2-1G9 protects macrophages from the cytotoxic effects of the Pseudomonas aeruginosa quorum sensing signalling molecule N-3-oxo-dodecanoyl-homoserine lactone. Mol Immunol 45:2710–2714CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Winter G, Griffiths AD, Hawkins RE, Hoogenboom HR (1994) Making antibodies by phage display technology. Annu Rev Immunol 12:433–455CrossRefPubMedGoogle Scholar
  6. 6.
    Bradbury AR, Sidhu S, Dübel S, McCafferty J (2011) Beyond natural antibodies: the power of in vitro display technologies. Nat Biotechnol 29:245–254CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Hermanson GT (2013) Bioconjugate techniques. Academic press, AmsterdamGoogle Scholar
  8. 8.
    Hoogenboom HR, Griffiths AD, Johnson KS, Chiswell DJ, Hudson P, Winter G (1991) Multi-subunit proteins on the surface of filamentous phage: methodologies for displaying antibody (Fab) heavy and light chains. Nucleic Acids Res 19:4133–4137CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Hayhurst A, Harris WJ (1999) Escherichia coli skp chaperone coexpression improves solubility and phage display of single-chain antibody fragments. Protein Expr Purif 15:336–343CrossRefPubMedGoogle Scholar
  10. 10.
    Pearson JP, Pesci EC, Iglewski BH (1997) Roles of Pseudomonas aeruginosa las and rhl quorum-sensing systems in control of elastase and rhamnolipid biosynthesis genes. J Bacteriol 179:5756–5767CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Papaioannou E, Wahjudi M, Nadal-Jimenez P, Koch G, Setroikromo R, Quax WJ (2009) Quorum-quenching acylase reduces the virulence of Pseudomonas aeruginosa in a Caenorhabditis elegans infection model. Antimicrob Agents Chemother 53:4891–4897CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  1. 1.Scottish Biologics FacilityUniversity of AberdeenAberdeenUK

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