Production of a human antibody fragment against the insulin-like growth factor І receptor as a fusion protein.

The aim of this study was to isolate single-chain variable fragments (scFvs) against human insulin-like growth factor I receptor (IGFIR) from a phage library displaying human scFvs. Isolated scFvs-displaying phages showed affinity for IGF-IR in comparison to the control. Expression of scFv proteins in Escherichia coli for further characterization, however, proved extremely difficult. Alternatively, the scFv protein was expressed as a fusion protein with a maltose-binding protein (MBP) that is a highly soluble E. coli protein. The MBPscFv fusion protein expressed in a soluble form in E. coli was purified to homogeneity by two-step affinity chromatography. The resulting MBP-scFv exhibited affinity for IGF-IR and structurally-related insulin receptor (IR). These results suggest both that MBPscFv fusion proteins are practical alternatives to isolating scFv proteins for further characterization and that successful isolation of human scFvs against a specific protein of interest requires vigorous screening in the early stages. Such screening is accomplished by using two independent screening methods such as measuring binding to IGF-IR but not to IR by ELISA or measuring competitive binding by IGF-I in addition to binding to IGF-IR alone.