MONITORING THE FORMATION OF SOLUBLE IMMUNE-COMPLEXES COMPOSED OF IDIOTYPE AND ANTIIDIOTYPE ANTIBODIES BY ELECTRON-MICROSCOPY

We have previously used immunoelectron microscopy (IEM) to generate a three-dimensional map of idiotypic (Id) and isotypic epitopes on the F ab arms of HGAC 39 (Roux et al., 1987, Proc. natn. Acad, Sci. U.S.A. 8 4, 4984-4988), a mouse IgG3 monoclonal antibody (Mab). In this report, we analyse the geometry of the various types of immune complexes form ed by the interaction of HGAC 39 with Mab directed against four mapped epitopes. Moreover, by sampling of reaction mixtures over time, we sh ow that the kinetics of each of the subpopulations of immune complexes , as defined by geometric configuration, can be determined. The data s how that for each antibody (Ab)-HGAC 39 combination the rate of immune complex formation was greatest during the first 1.5-3.5 min but that additional complexes formed through the remainder of the half hour ass ay period. As anticipated, complexes composed of even number units pre dominated (primarily dimers and tetramers) and most of these were in t he form of closed rings. The data also suggest that the location and o rientation of the epitopes on HGAC 39 to which the monoclonal antibodi es were bound has an influence on the types of immune complexes genera ted. Specifically we observed that those anti-idiotype Abs that bind t o the distal tip of Fab arms (i.e. in the CDR) are less likely to prod uce bivalently associated ringed dimers than antibodies that bind to e pitopes that are proximal to the CDR and that project laterally from t he surface of the Fab arms. These data are interpreted in terms of res trictions on hinge mediated flexibility and steric inhibition between adjacent Fab arms on HGAC 39.