Recombinant Genetic Libraries and Human Monoclonal Antibodies

  • Jarrett J. Adams
  • Bryce Nelson
  • Sachdev S. Sidhu
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1060)

Abstract

In order to comprehensively manipulate the human proteome we require a vast repertoire of pharmacological reagents. To address these needs we have developed repertoires of synthetic antibodies by phage display, where diversified oligonucleotides are used to modify the complementarity-determining regions (CDRs) of a human antigen-binding fragment (Fab) scaffold. As diversity is produced outside the confines of the mammalian immune system, synthetic antibody libraries allow us to bypass several limitations of hybridoma technology while improving the experimental parameters under which pharmacological reagents are produced. Here we describe the methodologies used to produce synthetic antibody libraries from a single human framework with diversity restricted to four CDRs. These synthetic repertoires can be extremely functional as they produce highly selective, high affinity Fabs to the majority of soluble human antigens. Finally we describe selection methodologies that allow us to overcome immuno-dominance in our selections to target a variety of epitopes per antigen. Together these methodologies allow us to produce human monoclonal antibodies to manipulate the human proteome.

Key words

Synthetic antibodies Phage display Synthetic libraries Protein engineering 
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Copyright information

© Springer Science+Business Media, LLC 2014

Authors and Affiliations

  • Jarrett J. Adams
    • 1
    • 2
    • 3
  • Bryce Nelson
    • 1
    • 2
    • 3
  • Sachdev S. Sidhu
    • 1
    • 2
    • 3
  1. 1.Terrence Donnelly Centre for Cellular and Biomolecular ResearchUniversity of TorontoTorontoCanada
  2. 2.Banting and Best Department of Medical ResearchUniversity of TorontoTorontoCanada
  3. 3.Department of Molecular GeneticsUniversity of TorontoTorontoCanada

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