Construction of a large naïve human phage-displayed Fab library through one-step cloning.
Antibody-based therapeutics is attracting more attention in the post-genome era, in contrast to a diminution in the initial high expectation for rapid development of gene-based therapeutic modalities. In support to the antibody-based therapeutics, the advent of recent technologies has made human antibody screening and production progressively more economic. Among those technologies, phage-display antibody library has been successfully applied in the antibody-based drug development both as fully human antibody sources and tools for antibody engineering. Building up a high-quality antibody library with a large library size and high diversity has been crucial for successful isolation of antibodies. Here we describe an efficient strategy for the construction of a large naïve phage-display human Fab library with one-step cloning. Optimization of each key step is extensively discussed and simplified protocols for library panning and Fab production are also described.
Zhu
Zhongyu
Z
NCI-Frederick, National Institutes of Health, Frederick, MD, USA.
Dimitrov
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D000941:Antigens / Q000276:immunology; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000096:biosynthesis; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000276:immunology; D008280:Magnetics; D008863:Microspheres; D008967:Molecular Biology / Q000379:methods*; D019151:Peptide Library; D012313:RNA / Q000302:isolation & purification
D000941:Antigens; D018076:DNA, Complementary; D007140:Immunoglobulin Fab Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D019151:Peptide Library; D012313:RNA