Insulin-like growth factor I/somatomedin-C: a rapid isolation procedure with FPLC.
Insulin-like growth factor I (IGF I)/somatomedin-C (SM-C) was purified from lyophilized human serum by acid-ethanol extraction. The extract was precipitated with acetone-ethanol. The precipitate was purified by Sephadex G-50 chromatography. The protein peak within a molecular weight range of 5000-10 000 was further purified with FPLC-reversed phase chromatography using a Pep RPC HR 5/5 column (Pharmacia) with a solvent system of acetonitrile (CH3CN) and 0.1% trifluoroacetic acid (TFA) in water. The purification of IGF I was monitored by radioimmunoassay for SM-C. Purity was established by analytical isoelectric focusing and by SDS polyacrylamide gel electrophoresis. Analytical isoelectric focusing showed one single protein band with an apparent pI of 8.3 +/- 0.1. SDS polyacrylamide gel electrophoresis showed also one single protein band with an apparent molecular weight of 7000. Biological activity was demonstrated by measuring the (3H)thymidine incorporation into DNA of cultured arterial smooth muscle cells.
D000096:Acetone; D001681:Biological Assay; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000096:biosynthesis; D000431:Ethanol; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000097:blood* / Q000494:pharmacology; D007525:Isoelectric Focusing; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism; D011863:Radioimmunoassay; D013002:Somatomedins / Q000097:blood*
D013002:Somatomedins; D000096:Acetone; D000431:Ethanol; D007334:Insulin-Like Growth Factor I; D004247:DNA