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Next-generation sequencing-guided identification and reconstruction of antibody CDR combinations from phage selection outputs.

Nucleic Acids Res.. 2019-03; 
BarretoKris, MaruthachalamBharathikumar V, HillWayne, HoganDaniel, SutherlandAshley R, KusalikAnthony, FongeHumphrey, DeCoteauJohn F, GeyerC Ro
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Recombinant Proteins … Page 4. 4 Nucleic Acids Research, 2019 (Constant Systems). Clarified supernatant was incubated with 200 μl of Protein-L resin (GenScript) for 1 h at 4 ? C. The Protein-L resin was collected by centrifugation and washed 5X with Protein-L binding buffer … Get A Quote

摘要

Next-generation sequencing (NGS) technologies have been employed in several phage display platforms for analyzing natural and synthetic antibody sequences and for identifying and reconstructing single-chain variable fragments (scFv) and antigen-binding fragments (Fab) not found by conventional ELISA screens. In this work, we developed an NGS-assisted antibody discovery platform by integrating phage-displayed, single-framework, synthetic Fab libraries. Due to limitations in attainable read and amplicon lengths, NGS analysis of Fab libraries and selection outputs is usually restricted to either VH or VL. Since this information alone is not sufficient for high-throughput reconstruction of Fabs, we develo... More

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