Interaction of a monoclonal IgE-specific antibody with cell-surface IgE-FcεRI: Characterization of equilibrium binding and secretory response

Richard G. Posner, Jodi M. Paar, Arieh Licht, Israel Pecht, Daniel H. Conrad, William S. Hlavacek

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Abstract

Aggregation of FcεRI, the high-affinity cell-surface receptor for IgE antibody, is required for degranulation of basophils and mast cells, but not all receptor aggregates elicit this cellular response. The stereochemical constraints on clusters of FcεRI that are able to signal cellular responses, such as degranulation, have yet to be fully defined. To improve our understanding of the properties of FcεRI aggregates that influence receptor signaling, we have studied the interaction of 23G3, a rat IgG IgE-specific monoclonal antibody, with IgE-FcεRI complexes on rat mucosal-type mast cells (RBL-2H3 line). We find that 23G3 is a potent secretagogue. This property and the structural features of 23G3 (two symmetrically arrayed IgE-specific binding sites) make 23G3 a potentially valuable reagent for investigating the relationship between FcεRI clustering and FcεRI-mediated signaling events. To develop a mathematical model of 23G3-induced aggregation of FcεRI, we used fluorimetry and flow cytometry to quantitatively monitor equilibrium binding of FITC-labeled 23G3 intact Ab and its Fab′ fragment to cell-surface IgE. The results indicate that IgE bound to FcεRI expresses two epitopes for 23G3 binding; that 23G3 binds IgE resident on the cell surface with negative cooperativity; and that 23G3 appears to induce mostly but not exclusively noncyclic dimeric aggregates of FcεRI. There is no simple relationship between receptor aggregation at equilibrium and the degranulation response. Further studies are needed to establish how 23G3-induced aggregation of IgE-FcεRI correlates with cellular responses.

Original languageEnglish (US)
Pages (from-to)11352-11360
Number of pages9
JournalBiochemistry
Volume43
Issue number35
DOIs
StatePublished - Sep 7 2004

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ASJC Scopus subject areas

  • Biochemistry

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