重组人碱性成纤维细胞生长因子的纯化及生物活性鉴定

Purification and bioactivity detection of recombinant human basic fibroblast growth factor

目的 :探讨重组人碱性成纤维细胞生长因子 (rhbFGF)的纯化工艺和活性鉴定 .方法 :将rhbFGF工程菌大量扩增后通过包涵体提取、复性、阳离子交换、凝胶过滤等技术进行纯化 .结果 :纯化后的rhbFGF ,相对分子质量为 17× 10 3,蛋白纯度为 95 %以上 ,比活为 1 7× 10 6 U/mg ,对NIH3T3细胞具有明显的促分裂活性 .结论 :通过复性和两步层析的纯化方法可获得高纯度、高回收率和高生物活性的rhbFGF 。

Aim: Purification and bioactivity detection of recombinant human basic fibroblast growth factor. Methods: rhbFGF was purified from rhbFGF producing BL21(DE3) bacteria. The inclusion bodies were first extracted and followed by cation-exchange chromatography and gel filtration. The bioactivities were detected by MTT assay. Results: rhbFGF was obtained with a purity above 95 percent, molecular weight is 1 7×10 3, and specific activity 1 7×10 6 U/mg. It enhances significantly the multiplication of 3T3 cells. Conclusion: rhbFGF of high purity and high bioactivity can be obtained by refolding of inclusion bodies and two steps of purification. The high yield and good quality of rhbFGF are important both for laboratory and the clinical application.