Interaction of a monoclonal antibody against hEGF with a receptor site forEGF

Epidermal growth factor (EGF) has been detected by radioimmunoassay (RIA) i n different body fluids such as serum, amniotic fluid, and urine. Human tum or tissues with EGF receptors (EGF-Rc) may be saturated with EGF, which may be of prognostic value. An RIA was envisaged to measure human epidermal gr owth factor (hEGF) levels using EGF Re as capture agent and a monoclonal an tibody anti-hEGF (MAb anti hEGF) labeled with (125)Iodine as a marker for t his binding. The purpose of this work was to study the feasibility of MAb a nti-hEGF to detect the receptor binding sites and to investigate the intera ction between MAb anti-hEGF and the EGF-Rc. Various binding experiments wer e performed to study possible interference and interactions in the complex MAb anti hEGF and the receptor. Affinity constants were determined by means of Scatchard plot analysis to interpret the complex stability challenged w ith other compounds for a better understanding of the interaction process. Binding constants were of the same order for all the ligands tested separat ely involving the EGF Rc, but were significantly higher (t = 15.7, P < 0.05 ) for hEGF in its binding to MAb anti-hEGF. It was possible with equilibriu m studies and competition experiments to evaluate the interaction of EGF an d MAb anti hEGF with the EGF receptor. This observation makes the MAb anti- hEGF a potential tracer for the quantitation of receptors in vitro, and pos sibly for the detection of membrane receptors on tumor cells in vivo. (C) 2 000 Elsevier Science Inc. All rights reserved.