Purification of a human urinary colony-stimulating factor.
Colony-stimulating factor (CSF), a protein required for the in vitro formation of colonies composed of granulocytes and/or macrophages, was isolated from the urine of anemic patients by using a seven-step procedure. The purified, homogeneous CSF had a specific activity of 1.9 X 10(8) U/absorbance unit at 280 nm (AU). This represents an overall purification of 25,330-fold and a total recovery of 3.8%. Upon iodination of the protein, the radioactivity migrated on sodium dodecyl sulfate (SDS) gel electrophoresis as a single peak with an apparent molecular weight of 46,000; reduction with mercaptoethanol caused dissociation to a single component of molecular weight 23,000. Only the dimer is active in stimulating colony formation. Urinary CSF stimulates formation of colonies comprising only macrophages in the mouse bone marrow cell culture assay. A neutralizing antibody raised against mouse L-cell CSF did not neutralize the activity of the urinary CSF but did bind it. This may indicate that the relative positions of antibody binding sites and the active sites are different in these two glycoproteins.
D002845:Chromatography / Q000379:methods; D002846:Chromatography, Affinity; D003115:Colony-Stimulating Factors / Q000276:immunology / Q000652:urine*; D006023:Glycoproteins / Q000302:isolation & purification; D006801:Humans; D008970:Molecular Weight
D003115:Colony-Stimulating Factors; D006023:Glycoproteins