A helper phage to improve single-chain antibody presentation in phage display (original) (raw)

Nature Biotechnology volume 19, pages 75–78 (2001)Cite this article

Abstract

We show here that the number of single-chain antibody fragments (scFv) presented on filamentous phage particles generated with antibody display phagemids can be increased by more than two orders of magnitude by using a newly developed helper phage (hyperphage). Hyperphage have a wild-type pIII phenotype and are therefore able to infect F+ Escherichia coli cells with high efficiency; however, their lack of a functional pIII gene means that the phagemid-encoded pIII–antibody fusion is the sole source of pIII in phage assembly. This results in an considerable increase in the fraction of phage particles carrying an antibody fragment on their surface. Antigen-binding activity was increased about 400-fold by enforced oligovalent antibody display on every phage particle. When used for packaging a universal human scFv library, hyperphage improved the specific enrichment factor obtained when panning on tetanus toxin. After two panning rounds, more than 50% of the phage were found to bind to the antigen, compared to 3% when conventional M13KO7 helper phage was used. Thus, hyperphage is particularly useful in stoichiometric situations, when there is little chance that a single phage will locate the desired antigen.

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Figure 1: The concept of hyperphage: a gene pIII-deleted helper phage with wild-type infection phenotype.

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Figure 2: Hyperphage increases presentation of scFv antibody fragments on filamentous phage.

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Figure 3: Hyperphage increases panning efficiency from a universal library.

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Acknowledgements

The financial support from the Heidelberger Akademie der Wissenschaften (Germany) is gratefully acknowledged. We wish to thank M. Little for support, M. Bechtel from Genzyme Virotech GmbH for supplying the tetanus toxin, and Olaf Broders for helping with the experimental work. We are grateful for E.K.F. Bautz for his continuous support and suggestions.

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Author notes

  1. Susanne Rondot and Joachim Koch: The first two authors contributed equally to this work.

Authors and Affiliations

  1. Molekulargenetisches Labor Prof. Raue, Friedrich-Ebert-Anlage 28, Heidelberg, 69117, Germany
    Susanne Rondot
  2. Forschungsstelle Hantaviren der Heidelberger Akademie der Wissenschaften, Im Neuenheimer Feld 230, Heidelberg, 69120, Germany
    Joachim Koch
  3. Abt. Genomanalyse, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, Heidelberg, 69120, Germany
    Frank Breitling
  4. Molekulare Genetik, Universität Heidelberg, Im Neuenheimer Feld 230, Heidelberg, 69120, Germany
    Stefan Dübel

Authors

  1. Susanne Rondot
  2. Joachim Koch
  3. Frank Breitling
  4. Stefan Dübel

Corresponding author

Correspondence toStefan Dübel.

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Rondot, S., Koch, J., Breitling, F. et al. A helper phage to improve single-chain antibody presentation in phage display.Nat Biotechnol 19, 75–78 (2001). https://doi.org/10.1038/83567

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