细胞实验检测蓖麻蚕蛹生物反应器表达的重组人表皮生长因子的生物学活性  

Expression of Recombinant Human Epidermal Growth Factor in Samia Cynthia Ricini Pupae Bioreactor and Its Biological Activity

作　　者：伍玉婷[1] 黄元姣[2] 雷丹青[2] 吴勇浒[1] 李牡艳[2] 

机构地区：[1]广西医科大学研究生学院,南宁市530021 [2]广西医科大学医学科学实验中心,南宁市530021

出　　处：《广西医学》2012年第6期673-676,共4页Guangxi Medical Journal

基　　金：广西科学研究与技术开发计划项目(10100019-20)

摘　　要：目的探讨蓖麻蚕蛹生物反应器中分离纯化获得的重组人表皮生长因子(rhEGF)蛋白质的生物学活性。方法采用Tricine-SDS-PAGE和免疫印迹法(Western blot)鉴定rhEGF,通过MTT和台盼蓝法检测rhEGF的细胞活性。结果 Tricine-SDS-PAGE、Western blot结果显示,蓖麻蚕蛹中分离纯化获得的rhEGF具有正确的分子量和免疫原性;MTT法显示,纯化的rhEGF具有酶促活性,且酶促活性优于rhEGF标准蛋白(P<0.05),其最高活性浓度为1.0μg/L,在0.0625～1.0μg/L浓度范围内具有剂量依赖性;rhEGF能够促进Balb/c3T3细胞的分裂增殖,效果优于rhEGF标准蛋白(P<0.05)。结论 AnpehEGF感染的蓖麻蚕蛹中分离纯化获得的rhEGF,具有类似天然hEGF的生物学活性,且效果优于rhEGF标准蛋白;rhEGF的细胞实验为进一步的动物实验提供了基础。Objective To study the biological activity of purified recombinant human epidermal growth factor （rhEGF） obtained from Samia cynthia ricini pupae bioreactor. Methods rhEGF was identify by Tricine-SDS-polyacrylamide gel electrophoresis （Tricine-SDS-PAGE） and Western blot, then methyl thiazolyl tetrazolium （MTI＇） assay and trypan blue staining technique were performed for the detection of the biological activity of balb/c3T3 cell. Results Successful expression of desired proteins could be observed by methods of Tricine-SDS-PAGE and Western blot. MTI＂ assay showed that the purified rhEGF caused the promotion of the enzyme activity in Balb/c3T3 cell in a dose-dependent manner ranged from 0.0625 to 1.0 μg/L. The best concentration in enzyme activity promoting was 1.0 μg/L, and its effect was stronger than that of standard rhEGF（P 〈 0.05）. The purified rhEGF could stimulate the proliferation of Balb/c3T3 ,and its effect was much more stronger in proliferation stimulating compared with the standard rhEGF（P 〈0.05）. Conclusion The purified rhEGF obtained from Samia cynthia ricini pupae bioreactor has similar biological activity with natural hEGF ,which has a better effect compared with standard rhEGF. This research provides a basis for the further animal test of rhEGF.

关 键 词：蓖麻蚕蛹生物反应器 重组人表皮生长因子 生物学活性 细胞实验 

分 类 号：Q78[生物学—分子生物学] Q74