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Identification of phage antibodies toward the Werner protein by selection on Western blots

✍ Scribed by Peter Ravn; Svend Kjær; Kristian Hobolt Jensen; Troels Wind; Kim Bak Jensen; Peter Kristensen; Robert M. Brosh; David K. Orren; Vilhelm A. Bohr; Brian F. C. Clark

Publisher: John Wiley and Sons
Year: 2000
Tongue: English
Weight: 336 KB
Volume: 21
Category: Article
ISSN: 0173-0835
DOI: 10.1002/(sici)1522-2683(20000201)21:3<509::aid-elps509>3.0.co;2-5

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✦ Synopsis

A procedure was established for selecting phage antibodies (phage-abs) from phage-displayed antibody repertoires by panning against proteins, separated by sodium dodecyl phosphate-polyacrylamide gel electrophoresis (SDS-PAGE) and electroblotted onto nitrocellulose membranes (Western blots). This immobilization strategy is applicable for secondary rounds of panning in selections against semipurified proteins, and directs the selection toward antibodies suitable as immunochemical reagents in Western blots. In model experiments, enrichment factors as high as 1.9x10(5) were obtained in a single round of panning. Furthermore, we demonstrate the application of this approach by selection of phage-abs recognizing the human Werner protein, which is defective in a premature aging syndrome.

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