Evidence for antibody activity against the receptor for IgE in a rabbit antiserum prepared against IgE-receptor complexes

D. H. Conrad, A. Froese, T. Ishizaka, K. Ishizaka

Research output: Contribution to journalArticle

Abstract

A rabbit antiserum prepared against rat IgE-receptor complexes was examined for evidence of antibody activity against receptors for IgE. The antibodies were specifically purified by absorption onto and subsequent elution from intact rat basophilic leukemia (RBL) cells and rendered specific for mast cells by absorption with mast cell-depleted rat peritoneal exudate cells. Surface-labeled RBL cells were extracted with Nonidet P-40 (NP-40) and the soluble extract was incubated with the purified antibodies; the components bound by the antibodies were analyzed on 10% SDS polyacrylamide gels. The major component bound by the antibodies had a relative mobility identical to that of the receptor for IgE. Confirmation that this component was indeed the receptor for IgE was shown in that this receptor was progressively removed from the NP-40 extract by increasing amounts of the purified antibodies. The ability of these purified antibodies to interact with the receptor was strongly inhibited when the receptor was complexed with IgE; in addition, the purified antibodies demonstrated little activity against purified IgE-receptor complexes. These results suggested that the antigenic site(s) interacting with the purified antibodies were either in or closely adjacent to the IgE-binding site on the receptor. In contrast, the original antiserum, after removal of anti-IgE antibodies by absorption with IgE covalently linked to Sepharose, demonstrated significant activity against the purified IgE-receptor complexes. This suggested that antibodies against sites on the receptor distant from the IgE-combining site were present in the original antiserum. These sites were probably buried in the membrane and thus lost in the antibody purification procedure.

Original languageEnglish (US)
Pages (from-to)507-512
Number of pages6
JournalJournal of Immunology
Volume120
Issue number2
StatePublished - Jan 1 1978

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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