The human combinatorial antibody library HuCAL GOLD combines diversification of all six CDRs according to the natural immune system with a novel display method for efficient selection of high-affinity antibodies
| Autor: | Corinna Löhning, Ingrid Pradel, Christine Rothe, Bodo Brocks, Ute Jager, Bernd Hubner, Yvonne Stark, Melanie Boss, Annemarie Honegger, Josef Prassler, Christian Frisch, Renate Bittlingmaier, Michael Bardroff, Tschimegma Bataa, Stefanie Urlinger, Margit Urban |
|---|---|
| Přispěvatelé: | University of Zurich, Rothe, C |
| Jazyk: | angličtina |
| Rok vydání: | 2008 |
| Předmět: |
Phage display
Protein Conformation Blotting Western Genetic Vectors Molecular Sequence Data Protein Data Bank (RCSB PDB) Antibody Affinity Immunoglobulin Variable Region Computational biology Biology Antibodies beta-Lactamases Immunoglobulin Fab Fragments 1315 Structural Biology Antibody Repertoire Antigen Structural Biology Peptide Library combinatorial antibody library 10019 Department of Biochemistry 1312 Molecular Biology Combinatorial Chemistry Techniques Humans Bacteriophages Amino Acid Sequence Cloning Molecular Peptide library Molecular Biology Peptide sequence Selection (genetic algorithm) human antibodies HuCAL GOLD Combinatorial chemistry Complementarity Determining Regions Genes Immune System biology.protein 570 Life sciences biology Immunoglobulin Light Chains Antibody phage display Immunoglobulin Heavy Chains CysDisplay |
| Popis: | This article describes the generation of the Human Combinatorial Antibody Library HuCAL GOLD. HuCAL GOLD is a synthetic human Fab library based on the HuCAL concept with all six complementarity-determining regions (CDRs) diversified according to the sequence and length variability of naturally rearranged human antibodies. The human antibody repertoire was analyzed in-depth, and individual CDR libraries were designed and generated for each CDR and each antibody family. Trinucleotide mixtures were used to synthesize the CDR libraries in order to ensure a high quality within HuCAL GOLD, and a beta-lactamase selection system was employed to eliminate frame-shifted clones after successive cloning of the CDR libraries. With these methods, a large, high-quality library with more than 10 billion functional Fab fragments was achieved. By using CysDisplay, the antibody fragments are displayed on the tip of the phage via a disulfide bridge between the phage coat protein pIII and the heavy chain of the antibody fragment. Efficient elution of specific phages is possible by adding reducing agents. HuCAL GOLD was challenged with a variety of different antigens and proved to be a reliable source of high-affinity human antibodies with best affinities in the picomolar range, thus functioning as an excellent source of antibodies for research, diagnostic, and therapeutic applications. Furthermore, the data presented in this article demonstrate that CysDisplay is a robust and broadly applicable display technology even for high-throughput applications. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|