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VHH characterization.Recombinant VHHs: Production, characterization and affinity.

Eric Chabrol1, Johann Stojko1, Alexandre Nicolas1

  • 1PEX Biotechnologies, Chimie, Biologie, Institut de Recherches Servier, 125 Chemin de Ronde, 78290, Croissy-sur-Seine, France.

Analytical Biochemistry
|November 3, 2019
PubMed
Summary

Researchers explored nanobodies (VHHs) as alternatives to antibodies for therapeutics. They developed methods to produce and analyze these small proteins, paving the way for complex VHH applications in areas like oncology.

Keywords:
AffinityCDCharacterizationDifferential scanning fluorimetryExpressionHydrogen/deuterium exchangeIon-mobilityMass spectrometryStructureSurface plasmon resonanceVHH

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Area of Science:

  • Biotechnology
  • Protein Engineering
  • Immunology

Background:

  • Antibodies are crucial therapeutics but have limitations.
  • Nanobodies (VHHs), derived from camelid heavy-chain antibodies, offer potential as smaller, more stable alternatives.
  • Developing robust methods for VHH production and characterization is essential for their therapeutic application.

Purpose of the Study:

  • To establish and validate analytical methodologies for nanobody (VHH) production and characterization.
  • To assess the feasibility of using VHHs as therapeutic agents.
  • To identify and overcome bottlenecks in VHH expression and purification.

Main Methods:

  • Expression of three standard VHHs (anti-albumin, anti-EGF receptor, anti-HER2) in various bacterial strains.
  • Application of a comprehensive suite of biophysical techniques, including spectroscopy and mass spectrometry.
  • Analysis of VHH structural features and the impact of disulfide bonds.

Main Results:

  • Successful expression and purification strategies were developed for homogeneous VHH preparations.
  • Established analytical methods confirmed VHH structural integrity and characteristics.
  • Identified key factors influencing VHH production and quality.

Conclusions:

  • The developed methodologies are suitable for producing and analyzing various nanobodies (VHHs).
  • These findings provide a foundation for developing more complex VHHs targeting specific diseases, particularly in oncology.
  • Nanobodies (VHHs) show promise as versatile therapeutic scaffolds.