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KMID : 0613820070170101321
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Journal of Life Science
2007 Volume.17 No. 10 p.1321 ~ p.1329
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Partial Purification and Quantification of Insulin-like Growth Factor-I from Red Deer Antler
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Gu Li Juan
Mo Eun-Kyung
Fang Zhe Ming
Sun Bai Shen
Zhu Xue Mei
Sung Chang-Keun
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Abstract
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Deer antler tissue contains the most rapidly growing bone in the animal kingdom. Thus, it is likely that growing antler tissue is a rich source of local paracrine bone-stimulating factors. Growth factors, at least the insulin-like growth factor (IGF), control the bone-remodelling process. In this study, we tried to isolate and purify IGF-I from fresh antler tissue by the routine isolation and purification of protein. The purification involved ammonium sulfate precipitation, DEAE-Sepharose CL-6B ion-exchange chromatography, CM-Sepharose CL-6B ion-exchange chromatography, and Sephadex G-50 chromatography. Purified fractions from each step were analyzed by high-performance liquid chromatography (HPLC), SDS polyacrylamide gel electrophoresis (SDS-PAGE), Dot-blot, and Western-blot methods. Furthermore, the quantification of partially purified IGF-I was calculated by enzyme-linked immunosorbent assays (ELISA) using antibody to human recombinant IGF-I. SDS-PAGE analysis of the final fraction yielded two molecular bands and the signal band was at 12 kDa on the Western-blot film. This purified IGF-I fraction showed a peak at retention time of eight min. The quantity of IGF-I in 20 g deer antler tissue as starting weight was calculated using a standard curve to be 2910 ng/ml, and total IGF-I amount is 0.291 g. The results show that IGF-I, which can be found in deer antler, can be partially purified and quantified by classic protein isolation methods.
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KEYWORD
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Deer antler, insulin-like growth factor (IGF), DEAE-Sepharose CL-6B, CM-Sepharose CL-6B, Sephadex G-50, HPLC, ELISA
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