Generation of Rabbit Monoclonal Antibody Fragments from a Combinatorial Phage Display Library and Their Production in the Yeast Pichia pastoris
- 1 March 1995
- journal article
- research article
- Published by Springer Nature in Nature Biotechnology
- Vol. 13 (3) , 255-260
- https://doi.org/10.1038/nbt0395-255
Abstract
We have applied the combinatorial immunoglobulin library and phage display technologies to generate monoclonal rabbit single-chain Fv (scFv) antibody fragments specific for recombinant human leukemia inhibitory factor (rhLIF). The B cell immunoglobulin repertoire of an immunized rabbit was immortalized by the combinatorial cloning of the rearranged variable domains of light (V-I) and heavy (V-H) chains. Affinity selection of the library displaying the rabbit antibody domains on the phage surface resulted in the isolation of phage encoding scFv antibodies which specifically bind to the antigen, We utilized the methylotrophic yeast Pichia pastoris for high level secretion of soluble and functional scFv antibody fragment. More than 100 mg/L of pure and functional rabbit anti-rhLIF scFv antibody was obtained directly from the P. pastoris culture supernatant by one-step affinity chromatography.Keywords
This publication has 38 references indexed in Scilit:
- Making Antibodies by Phage Display TechnologyAnnual Review of Immunology, 1994
- Bacterial expression of immunoglobulin fragmentsCurrent Opinion in Immunology, 1993
- Selection of phage antibodies by binding affinityJournal of Molecular Biology, 1992
- Restricted VH Gene Usage and Generation of Antibody Diversity in RabbitAnnual Review of Immunology, 1992
- By-passing immunizationJournal of Molecular Biology, 1991
- Production of mouse epidermal growth factor in yeast: high-level secretion using Pichia pastoris strains containing multiple gene copiesGene, 1991
- Making antibody fragments using phage display librariesNature, 1991
- Phage antibodies: filamentous phage displaying antibody variable domainsNature, 1990
- Molecular basis of the allelic inheritance of rabbit immunoglobulin VH allotypes: Implications for the generation of antibody diversityCell, 1990
- Diversity in the rabbit immunoglobulin kappa chain variable regions is amplified by nucleotide deletions and insertions at the V-J junctionCell, 1983