Preparation of phage antibodies to the ED-A domain of human fibronectin

The fibronectin (FN) isoform containing the alternative spliced ED-A domain is much more expressed in fetal, tumoral, and regenerating tissues than in normal adult tissues. The ED-A containing FN is up-regulated by numerous cytokines, such as TGF-β, and, although in normal adult liver the ED-A domain is undetectable, in regenerating rat liver the expression of ED- A is increased and mediates the conversion of fat storing cells to myofibroblasts. Here we describe the selection from a phage display library and the characterization of human antibody fragments directed against the EDA sequence of FN. As they can be easily radiolabeled with ³²P, these antibodies are very highly sensitive reagents for the determination of ED-A levels in tissues and biological fluids; in fact, use of these scFv induced a more than 10-fold increase in sensitivity with respect to the murine monoclonal IST-9. The possibility of preparing a range of human engineered antibodies should facilitate the development of antibody reagents with suitable pharmacokinetics, valency, functional affinity, and effector functions and that could be useful for clinical purposes.