Purification and characterization of an endothelial cell growth factor from serum-free culture medium of human diploid fibroblast cells.
Serum-free culture supernatants of human embryo fibroblast cells contain endothelial cell growth factor (f-ECGF) which supports the serial propagation of human umbilical vein endothelial cells in the serum-free culture medium (medium A). This growth-stimulating activity has been partially purified from serum-free culture-conditioned medium. The stability of the activity to acid (pH 4.0-4.5) was utilized for the first step in purification. f-ECGF had a high affinity to heparin-Sepharose CL-6B, and was isolated by the methods of heparin affinity, of ion-exchange and gel filtration chromatography from the serum-free culture-conditioned medium preparation. The purified f-ECGF had an isoelectric point in the pH range 4.5-6, and a molecular weight of approx. 30 kDa determined by either gel filtration or SDS-polyacrylamide gradient gel electrophoresis. The f-ECGF has high affinity for concanavalin A column, and the activity was partially eluted from the column with ethylene glycol and alpha-methylmannose. The results indicate that f-ECGF is an acidic-glyco-protein with heterogeneous sugar chain(s).
D002460:Cell Line; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003470:Culture Media; D004171:Diploidy; D016228:Endothelial Growth Factors; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D006133:Growth Substances / Q000302:isolation & purification* / Q000502:physiology; D006801:Humans; D007525:Isoelectric Focusing; D008168:Lung / Q000196:embryology; D008970:Molecular Weight; D011247:Pregnancy; D014471:Umbilical Veins / Q000166:cytology
D003470:Culture Media; D016228:Endothelial Growth Factors; D006133:Growth Substances