Binding of rat IgE with the subcellular components of normal rat mast cells

Wolfgang König, Kimishige Ishizaka

Research output: Contribution to journalArticlepeer-review

Abstract

Attempts were made to obtain subcellular components of rat mast cells which contain receptors for IgE. Rat peritoneal cells were treated with 125I-labeled rat IgE to label mast cells, and disrupted by ultrasonication. More than 90% of cell-bound IgE was recovered in the 20,000 g supernatant fraction which had the ability to block passive cutaneous anaphylaxis with mouse and rat reaginic antibodies. Gel-filtration of the supernatant fraction through a Sepharose 6B column showed that both radioactivity and PCA blocking activity were associated with subcellular components which were eluted in the void volume. When the 20,000 g supernatant fraction of normal peritoneal cells was incubated with 125I-rat IgE, the protein combined with the subcellular components. Incubation of the 20,000 g supernatant with an IgE-rich rat serum inhibited the binding of 125I-IgE with the components, whereas normal rat serum failed to do so. When the sepharose void volume fraction was obtained from purified mast cells and labeled with 125I, approximately 25% of radioactive material was coprecipitated with IgE-anti-IgE complexes. Evidence was obtained that subcellular components of macrophages were not coprecipitated with the antigen-antibody complexes. These results indicated that the components eluted in the void volume fraction contained receptors for IgE. In another experiment, the 20,000 g supernatant fraction was obtained from rat peritoneal cells or purified mast cells whose membrane was labeled with 125I, and fractioned by gel filtration followed by DEAE cellulose chromatography. More than 2 3 of the radioactivity and PCA-blocking activity associated with the sepharose void volume fraction were recovered in the 3 M NaCl fraction which contained 12-15% of total protein. When the 3 M fraction obtained from purified mast cells was labeled with 125I and incubated with an IgE-rich serum, a significant amount of radioactive material in the fraction was precipitated with IgE-anti-IgE complexes. The results indicate that blocking of the PCA reaction by this fraction is due to the presence of receptors for IgE.

Original languageEnglish (US)
Pages (from-to)345-353
Number of pages9
JournalImmunochemistry
Volume13
Issue number4
DOIs
StatePublished - Apr 1976

ASJC Scopus subject areas

  • Medicine(all)

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