Abstract：
:Recently the analytical power of the latest high throughput next generation DNA sequencing platforms has been used to analyse phage that have been selected from the panning of large combinatorial libraries displaying either peptide or antibody ligands. This process, commonly referred to as next generation phage display (NGPD), allows the researcher to determine the identity of specific phage that are being enriched against an antigen target by analysis of the DNA sequence encoding the displayed ligand. This method bypasses several steps in conventional phage panning that include laborious colony picking and functional ligand screening. A downside of this approach is that the only output from such experiments is the DNA sequence information of such enriched phage particles. In the case of peptides, the peptide sequence can be synthesised directly and used for further screening; however this is more difficult with larger antibody fragments such as ScFvs. In the case of ScFvs, their coding sequence would have to be fully elucidated, synthesised and re-cloned before expression. We describe here the application of an inverse PCR-ligation methodology that enables the specific recovery of ScFvs of interest from enriched sub-libraries of phage clones. Phagemid particles are recovered using sequence information derived from their unique heavy chain CDR3/FR4 domains and specific clones can be recovered irrespective of CDR3 size and at levels of abundance that would be refractory to their discovery during conventional phage panning and screening.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Spiliotopoulos A,Owen JP,Maddison BC,Dreveny I,Rees HC,Gough KCdoi
10.1016/j.jim.2015.03.005subject
Has Abstractpub_date
2015-05-01 00:00:00pages
50-5eissn
0022-1759issn
1872-7905pii
S0022-1759(15)00069-1journal_volume
420pub_type
杂志文章abstract：:Drug-specific monoclonal antibodies (MAbs) were produced against the very small drug hapten (162.15 Da), 5-benzimidazolecarboxylic acid, an analogue of 2-(4-Thiazolyl)benzimidazole (TBZ) but lacking the thiol group. TBZ is widely used as a broad-spectrum anthelmintic in various animal species and humans and also as a ...
journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
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journal_title：Journal of immunological methods
pub_type： 杂志文章
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