IMMUNOCHEMICAL STUDY OF HUMAN-IMMUNOGLOBULIN-G FC REGION

Fifteen murine monoclonal antibodies (mAb) were produced that reacted with conformational epitopes present on the Fc portion of all human Ig G subclasses (PAN-Fc). Inhibition and sandwich ELISA were used to eluc idate overlapping and distinct epitopes. The epitopes were aligned to form a continuous ''chain'' of overlapping determinants from the C(H)2 -C(H)3 interface to the direction of hinge. The five more hinge-proxim al epitopes demonstrated high lability both in competitive and sandwic h assays, being completely or partially destroyed when any of these 15 other mAb bound the IgG first. Heat-inactivation of IgG caused full d isruption of these epitopes. In contrast, epitopes situated at the opp osite distal of the ''chain'' were more stable and mAb binding could o nly be affected by occupying an overlapping epitope. Under heat-inacti vation these epitopes were affected, but not completely destroyed. Hum an IgG class anti-DNA antoantibodies were bound to insolubilized dsDNA and their reaction with PAN-Fc mAb was studied. mAb titration plots o n IgG and dsDNA-IgG were compared. Five epitopes proved to be altered by antigen (dsDNA) binding. Two of these were the labile hinge-proxima l epitopes and the other three were situated near the C(H)2-C(H)3 inte rface. Cross-reactivity of mAb with xenogeneic IgG was also studied. A n ''epitope map'' of the crystallographic model of human IgG Fc portio n drawn was based on these experimental data and printed matter, conce rning the location of subclass-specific amino acids and homology regio ns of humans and animal IgG.