Back to Search
Start Over
Selection of affibody molecules to the ligand-binding site of the insulin-like growth factor-1 receptor.
- Source :
-
Biotechnology and applied biochemistry [Biotechnol Appl Biochem] 2010 Feb 25; Vol. 55 (2), pp. 99-109. Date of Electronic Publication: 2010 Feb 25. - Publication Year :
- 2010
-
Abstract
- Affibody molecules binding to the site of hormone interaction in IGF-1R (insulin-like growth factor-1 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 IGF-1 (insulin-like growth factor-1). 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-1 competition during elution, in addition to affibody molecules generated in the non-competitive selections, could compete with IGF-1 for binding to the receptor. The affinities of the isolated variants to IGF-1R-overexpressing MCF-7 cells were determined and ranged from high nanomolar to 2.3 nM. The most promising variant, Z4:40, was shown to recognize IGF-1R efficiently in several different contexts: in analyses based on flow cytometry, fluorescence microscopy and receptor pull-down from cell extracts. In addition, when Z4:40 was added to the medium of MCF-7 cells that were dependent on IGF-1 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-1R status, as well as applications of affibody-based reagents for therapy, are discussed.
- Subjects :
- Animals
Antibodies, Monoclonal genetics
Antibodies, Monoclonal immunology
Cell Line, Tumor
Enzyme-Linked Immunosorbent Assay
Humans
Mice
Receptor, IGF Type 1 immunology
Single-Chain Antibodies genetics
Single-Chain Antibodies immunology
Antibodies, Monoclonal chemistry
Receptor, IGF Type 1 chemistry
Single-Chain Antibodies chemistry
Subjects
Details
- Language :
- English
- ISSN :
- 1470-8744
- Volume :
- 55
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- Biotechnology and applied biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 20088825
- Full Text :
- https://doi.org/10.1042/BA20090226