Purification of a granulocyte colony-stimulating factor from the conditioned medium of a subclone of human bladder carcinoma cell line 5637, HTB9.
A colony-stimulating factor (CSF) has been purified to homogeneity from the conditioned medium (CM) of a subclone, designated HTB9, of human bladder carcinoma cell line 5637. HTB9 cells were successfully cultured on micro-carrier beads at low-serum concentration, and the resulting CM (HTB9-CM) was concentrated by ultrafiltration. Purification procedures consisted of anion-exchange column chromatography, gel-filtration column chromatography, and reverse-phase column chromatography. The finally purified protein possessed a specific activity more than 10(8) U/mg protein for day-7 CFU-GM colony formation and exhibited a single band representing a molecular weight of 17,000 upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Biological activity was apparently specific for a neutrophilic granulocyte lineage of human non-phagocytic bone-marrow cells in vitro, and antiserum raised against this purified protein completely inhibited the activity of recombinant granulocyte colony-stimulating factor.
First Department of Internal Medicine, Faculty of Medicine, Kyushu University, Fukuoka, Japan.
E
E
Morioka
S
S
Taniguchi
S
S
Okamura
T
T
Shibuya
Y
Y
Niho
D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002999:Clone Cells; D003470:Culture Media; D016179:Granulocyte Colony-Stimulating Factor / Q000302:isolation & purification*; D006801:Humans; D007106:Immune Sera; D014407:Tumor Cells, Cultured; D001749:Urinary Bladder Neoplasms / Q000737:chemistry*
D003470:Culture Media; D007106:Immune Sera; D016179:Granulocyte Colony-Stimulating Factor