酸性成纤维细胞生长因子的分离纯化及活性鉴定

Purification of aFGF from Bovine Brain and Identification of Its Biological Activity

目的 :从牛脑中分离纯化酸性成纤维细胞生长因子 (aFGF)并鉴定生物活性。方法 :根据DenisGOSPODAROW ICZ的方法 ,新鲜牛脑匀浆 ,三步硫酸铵盐析 ,最后沉淀物溶解透析后予离子交换层析及琼脂糖凝胶亲和层析。促 3T3细胞DNA合成率鉴定生物活性。结果 :用 1.0mol/LNaCl的缓冲液洗脱得到分子量为 136 0 0的多肽 ,得率为 6 10 μg/kg牛脑。氨基酸组成分析与已知的aFGF相同。促 3T3细胞DNA合成的最低浓度为 0 .5ng/ml,最大效应浓度 5 0ng/ml,ED50 值为 8ng/ml。结论

Objective: To purify aFGF from bovine brain and to determine its mitogenic activity. Methods: aFGF was purified from bovine brain by the Denis GOSPODAROWICZ assay. Fresh bovine brain was homogenized and extracted by three different concentrations of ammonium sulfate.The precipitation was followed by cation exchange and sepharose affinity chromatography. The bioassay was based on the stimulation of DNA synthesis in 3T3 cells. Results: A polypeptide is eluted from 1.0 mol/L NaCl, with a molecular weight of 13 600, a productive rate of 610 μg/kg from bovine brain. Its amino acid composition and biological characteristics showed the polypeptide was identical to aFGF. It was active in stimulating DNA synthesis in 3T3 cells at concentration as low as 0.5 ng/ml. Maximal effect was achieved at 50 ng/ml, with ED 50 of 8 ng/ml. Conclusion: The results demonstrate that aFGF is a potent mitogenic factor.