ArticleNucleic acids research2009
Bicistronic DNA display for in vitro selection of Fab fragments.
Article in Nucleic acids research, 2009. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed, 20 citations in OpenAlex.
- Cell-Free Gene Expression: Methods and Applications.Chemical reviews · 2025Review
- Review
- Conceptual and methodological advances in cell-free directed evolution.Current opinion in structural biology · 2015Review
- A Novel Dual Expression Platform for High Throughput Functional Screening of Phage Libraries in Product like Format.PloS one · 2015Article
- In vitro Fab display: a cell-free system for IgG discovery.Protein engineering, design & selection : PEDS · 2014Article
- Selection of a T7 promoter mutant with enhanced in vitro activity by a novel multi-copy bead display approach for in vitro evolution.Nucleic acids research · 2013Article
- In vitro evolution of enzymes.Methods in molecular biology (Clifton, N.J.) · 2013Article
- DNA display selection of peptide ligands for a full-length human G protein-coupled receptor on CHO-K1 cells.PloS one · 2012Article
- In vitro selection of fab fragments by mRNA display and gene-linking emulsion PCR.Journal of nucleic acids · 2012Article
- A Concept for Selection of Codon-Suppressor tRNAs Based on Read-Through Ribosome Display in an In Vitro Compartmentalized Cell-Free Translation System.Journal of nucleic acids · 2012Article
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
In vitro display methods are superior tools for obtaining monoclonal antibodies. Although totally in vitro display methods, such as ribosome display and mRNA display, have the advantages of larger library sizes and quicker selection procedures compared with phage display, their applications have been limited to single-chain Fvs due to the requirement for linking of the mRNA and the nascent protein on the ribosome. Here we describe a different type of totally in vitro method, DNA display, that is applicable to heterodimeric Fab fragments: in vitro compartmentalization in water-in-oil emulsions allows the linking of an oligomeric protein and its encoding DNA with multiple ORFs. Since previously used emulsions impaired the synthesis of functional Fab fragments, we modified conditions for preparing emulsions, and identified conditions under which it was possible to enrich Fab fragments 10(6)-fold per three rounds of affinity selection. Furthermore, we confirmed that genes encoding stable Fab fragments could be selected from a Fab fragment library with a randomized hydrophobic core in the constant region by applying heat treatment as a selection pressure. Since this method has all advantages of both phage display and totally in vitro display, it represents a new option for many applications using display methods.
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