Solubilization and assay of a colony‐stimulating factor receptor from murine macrophages

Yee‐Guide ‐G Yeung, Paul T. Jubinsky, E. Richard Stanley

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

The colony‐stimulating factor, CSF‐1, selectively stimulates the survival, proliferation, and differentiation of mononuclear phagocytes. The solubilization, assay, and characteristics of the CSF‐1 receptor from the J774.2 murine macrophage cell line are described. The recovery of cell‐surface receptor in the postnuclear supernatant membrane fraction of hypotonically disrupted cells was 76%. Recovery of the ligand binding activity of the receptor after solubilization of this fraction with 1% Triton X‐100 was ∼ 150%. The binding of 125I‐CSF‐1 to intact cells and membrane preparations was consistent with the existence of a single class of high‐affinity receptor sites. In contrast, the equilibrium binding of 125I‐CSF‐1 to the solubilized postnuclear fraction indicated the existence of two distinct classes of binding site (apparent Kds 0.15 nM and 10 nM). A rapid assay was developed for the high‐affinity sites, which were shown to be associated with the CSF‐1 receptor. The function of the low‐affinity sites, which have not been demonstrated on intact cells or cell membranes and which are 13 times more abundant than the high‐affinity sites, is unknown. The solubilized high‐affinity receptor‐CSF‐1 complex was stable on storage at 0°C and −70°C but dissociated at 37°C. Dissociation also occurred at 0°C in buffers of low pH (4.0) or high ionic strength (0.7 M NaCl).

Original languageEnglish (US)
Pages (from-to)259-269
Number of pages11
JournalJournal of Cellular Biochemistry
Volume31
Issue number4
DOIs
Publication statusPublished - 1986

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Keywords

  • CSF‐1
  • CSF‐1 receptor
  • colony‐stimulating factor
  • membrane proteins
  • mononuclear phagocytes
  • murine macrophages

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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