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Miniantibodies

  • Protocol
Antibody Engineering

Part of the book series: Springer Protocols Handbooks ((SPH))

Abstract

Recombinant antibodies have become a standard component of research, diagnostics, and therapy. In the development of recombinant antibodies – irrespective of the final format – a monovalent construct is virtually always the first protein to be tested. This is due to the fact that essentially all selection systems use such formats, and that the periplasmic production of scFv and Fab fragments has now become standard. Nonetheless, some tasks require an increase of the avidity to the respective antigens, antibody, as well as the fragment size. A convenient way to rather quickly achieve both is by fusing a hinge region followed by a dimerizing or oligomerizing structure to the C-terminus of the antibody fragment, creating the so-called “miniantibodies”. Compared to other available bivalent or bispecific formats, miniantibodies distinguish themselves by their rotational freedom and flexibility, being similar to full-length antibodies. This protocol describes the modular conversion of scFv fragments into miniantibodies, resulting in a final multimerized structure with two or four binding sites, and we present different self-associating domains suitable for this task. Additionally, we also provide information on their production and discuss how to improve the yield of soluble antibody fragments.

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Abbreviations

IPTG:

Isopropylthiogalactoside

PBS:

Phosphate buffered saline

scFv:

Single-chain Fv fragment

tet:

Tetracycline

LB:

Luria-Bertani media

SB:

Super broth media

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Acknowledgements

This chapter is based on the original work of Peter Pack, Jörg Willuda and Susanne Kubetzko, with subsequent contributions from Kerstin Blank and Barbara Klinger.

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Authors and Affiliations

  1. Biochemisches Institut, Universität Zürich, Winterthurerstrasse 190, 8057, Zürich, Switzerland

    Jonas V. Schaefer, Peter Lindner & Andreas Plückthun

Authors
  1. Jonas V. Schaefer
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  2. Peter Lindner
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  3. Andreas Plückthun
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Corresponding author

Correspondence to Andreas Plückthun .

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Editors and Affiliations

  1. (Biomedical Engineering) Institut für Zellbiologie und Immunologie, Universität Stuttgart, Allmandring 31, 70569, Stuttgart, Germany

    Roland Kontermann

  2. Technische Universität Braunschweig, Institut für Biochemie und Biotechnologie, Spielmannstraße 7, 38106, Braunschweig, Germany

    Stefan Dübel

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Schaefer, J.V., Lindner, P., Plückthun, A. (2010). Miniantibodies. In: Kontermann, R., Dübel, S. (eds) Antibody Engineering. Springer Protocols Handbooks. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-01147-4_7

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  • DOI: https://doi.org/10.1007/978-3-642-01147-4_7

  • Publisher Name: Springer, Berlin, Heidelberg

  • Print ISBN: 978-3-642-01146-7

  • Online ISBN: 978-3-642-01147-4

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