In the present study, two insulin-like growth factor-binding protein 7 (IGFBP7) C-terminal-peptides were expressed as fusion proteins to bacterial verotoxin pentamerization domain as shown by Western blotting, ELISA and mass spectroscopy. Both in vivo-biotinylated recombinant products were purified from bacterial lysates by IMAC and used directly for panning along with the recombinant IGFBP7 protein using the LAC-M Camelidae naïve single domain antibody (sdAb) library. Target-specific sdAbs to both parental protein and peptide fusions were identified by phage ELISA. Twelve different clones were isolated by phage-ELISA screening and their sdAb genes were sequenced. Soluble sdAbs and their pentameric formats were expressed in TG1 E. coli, purified by IMAC and characterized by ELISA and SPR. Several sdAbs are currently under study, however anti-IGFBP7 (P12/M12) was extensively characterized and exhibited promising anti-tumorigenic effect on PANC-1 cell lines by blocking IGFBP7 promoting activity. This study provides the basis for developing a novel imaging/therapeutic reagent for targeting and treating brain tumor angiogenesis in early stages of tumorogenesis and can also be used as a molecular tool to monitor the degree of angiogenesis in gliomas which may help to improve the clinical management of brain tumors.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/23523 |
| Date | January 2012 |
| Creators | Alturki, Norah |
| Contributors | Tanha, Jamshid |
| Publisher | Université d'Ottawa / University of Ottawa |
| Source Sets | Université d’Ottawa |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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