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Selection of affibody molecules to the ligand-binding site of the insulin-like growth factor-1 receptor
KTH, School of Biotechnology (BIO), Molecular Biotechnology (closed 20130101).
KTH, School of Biotechnology (BIO), Molecular Biotechnology (closed 20130101).ORCID iD: 0000-0001-7034-0850
KTH, School of Biotechnology (BIO), Molecular Biotechnology (closed 20130101).
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2010 (English)In: Biotechnology and applied biochemistry, ISSN 0885-4513, E-ISSN 1470-8744, Vol. 55, 99-109 p.Article in journal (Refereed) Published
Abstract [en]

Affibody molecules binding to the site of hormone interaction in IGF-IR (insulin-like growth factor-I receptor) were successfully selected by phage-display technology employing a competitive-elution strategy during biopanning, whereby release of receptor-bound phagemids was accomplished by competition with IGFI (insulin-like growth factor-I). In non-competitive selections, the elution of receptor-bound phagemids was performed by imidazole or low-pH incubation, which also resulted in the isolation of affibody molecules that could bind to the receptor. An ELISA-based assay showed that the affibody molecules generated by IGF-I competition during elution, in addition to affibody molecules generated in the noncompetitive selections, could compete with IGF-I for binding to the receptor. The affinities of the isolated variants to IGF-IR-overexpressing MCF-7 cells were determined and ranged from high nanomolar to 2.3 nM. The most promising variant, Z(4;40), was shown to recognize IGF- IR efficiently in several different contexts: in analyses based on flow cytometry, fluorescence microscopy and receptor pull-down from cell extracts. In addition, when Z, was added to the medium of MCF-7 cells that were dependent on IGF-I for efficient growth, it was found to have a dose-dependent growth-inhibitory effect on the cells. Applications of affibody-based reagents for quantitative and qualitative analyses of IGF- I R status, as well as applications of affibody-based reagents for therapy, are discussed.

Place, publisher, year, edition, pages
2010. Vol. 55, 99-109 p.
Keyword [en]
affibody molecule, competitive elution, insulin-like growth factor-1, receptor (IGF-IR), phage display, human-breast-cancer, monoclonal-antibody, carcinoma cells, proteins, domain, expression, libraries, scaffold
National Category
Industrial Biotechnology
Identifiers
URN: urn:nbn:se:kth:diva-19387DOI: 10.1042/ba20090226ISI: 000276539400005Scopus ID: 2-s2.0-77954828431OAI: oai:DiVA.org:kth-19387DiVA: diva2:337434
Funder
Swedish Research Council, 621-20055134
Note

QC 20100525

Available from: 2010-08-05 Created: 2010-08-05 Last updated: 2017-12-12Bibliographically approved

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Lundberg, EmmaGräslund, Torbjörn

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