Factors regulating macrophage production and growth. Purification and some properties of the colony stimulating factor from medium conditioned by mouse L cells

E. R. Stanley, P. M. Heard

Research output: Contribution to journalArticle

451 Scopus citations

Abstract

A five-step purification procedure involving concentration, chromatography on DEAE-cellulose, Sephadex G-200 concanavalin A/Sepharose and gradient gel electrophoresis has been developed for a factor (colony stimulating factor, CSF) produced by cultured mouse L cells. CSF stimulates granulocyte-macrophage production from single hemopoietic progenitor cells and shares identity with macrophage growth factor. It was purified from a higher molecular weight, concanavalin A-adherent fraction that represented approximately 70% of the total colony stimulating activity of serum-free mouse L cell-conditioned medium. The purified material had a specific biological activity of 1.62x108 units/mg of protein and stimulated maximum colony formation in the standard assay system at a concentration of 10-11 M. On both standard and sodium dodecyl sulfate-polyacrylamide gel disc electrophoresis it gave a single band of protein that co-electrophoresed with a single band of biological activity. It behaved heterogeneously on isoelectric focusing in polyacrylamide gels, yielding several bands of protein in the pH range 3.7 to 4.9, each of which was associated with biological activity. The protein and biological activity of the purified material were shown to combine with components in antiserum raised against the penultimate purification stage at equivalent antiserum concentrations. The purified CSF is a glycoprotein with a molecular weight of approximately 70,000, composed of two similarly charged polypeptide chains of molecular weight approximately 35,000, covalently bound through disulfide bonds.

Original languageEnglish (US)
Pages (from-to)4305-4312
Number of pages8
JournalJournal of Biological Chemistry
Volume252
Issue number12
StatePublished - Dec 1 1977

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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