bFGF生物蛋白海绵体外生物学活性检测方法的建立及其稳定性评价

J4 ›› 2011, Vol. 37 ›› Issue (1): 171-174.

• 基础研究 • 上一篇下一篇

bFGF生物蛋白海绵体外生物学活性检测方法的建立及其稳定性评价

焦悦^1|2|王晓杰^1|3|张睿^1|2|蔡敏倩¹|李婷婷¹|高丽昌¹|王珊珊⁴|李海燕¹|李校堃^1|3

1.吉林农业大学生物反应器与药物开发教育部工程研究中心,吉林长春 130118； 2.吉林农业大学中药材学院,吉林长春 130118； 3.温州医学院药学院 |浙江省生物技术制药工程重点实验室,浙江温州 325000；4.哈尔滨沛奇隆生物制药有限公司,黑龙江哈尔滨 150090

收稿日期:2010-04-27 出版日期:2011-01-28 发布日期:2011-01-28
通讯作者: 李校堃 (Tel:0431-84533348,E-mail:xiaokunli@163.net); 李海燕 (Tel:0431-84533347,E-mail：hyli99@163.com) E-mail:xiaokunli@163.net；hyli99@163.com
作者简介:焦悦（1985-）|女|吉林省辽源市人|在读农学硕士|主要从事基因工程制药方面的研究
基金资助:
国家863计划项目资助课题（2007AA100503）；教育部科技创新工程重大项目-培养资金项目资助课题;
吉林省长春市科技特派员行动计划项目资助课题（09KT03）；吉林省科技厅科技发展计划项目资助课题（20070922）

Establishment of method for detection of biological activity of |biological |protein sponge of |basic fibroblast growth factor |and stability evaluation

JIAO Yue^1|2|WANG Xiao-jie^1|3|ZHANG Rui^1|2|CAI Min-qian¹|LI Ting-ting¹|GAO Li-chang¹|WANG Shan-shan⁴|LI Hai-yan¹|LI Xiao-kun^1|3

1.Engineering Research Center of Bioreactor and Pharmaceutical Development,Ministry of Education,Jilin Agricultural Univerity,Changchun 130118,China;2. College of Traditional Chinese Medicine,Jilin Agricultural Univerity,Changchun 130118,China;3.Key Laboratory of Biotechnology Pharmaceutical Engineering, |Zhejiang Province,Department of Pharmacy, Wenzhou Medical College,Wenzhou 325000,China；4.Harbin Peiqilong Biological Pharmaceutical Co.,Ltd,Harbin 150090,China

Received:2010-04-27 Online:2011-01-28 Published:2011-01-28

摘要/Abstract

摘要：

目的：建立碱性成纤维细胞生长因子（bFGF）生物蛋白海绵体外生物学活性测定方法观察bFGF生物蛋白海绵常温放置的稳定性。方法：bFGF生物蛋白海绵供试品融于稀释液中，离心后获得浸提液，以bFGF标准品为对照，与NIH3T3细胞共培养，MTT法检测bFGF生物蛋白海绵浸提液对NIH3T3细胞的促增值作用，用bFGF标准品计算bFGF生物蛋白海绵的相对百分效价，对该方法进行验证；以常温储存2、3和4年的bFGF生物蛋白海绵为供试品，以低温储存30 d的bFGF生物蛋白海绵为标准对照，以不含bFGF生物蛋白海绵和bFGF的培养液为空白对照，测定各组生物学活性效价，比较不同储存时间bFGF生物蛋白海绵的体外生物学活性效价的稳定性。结果：bFGF生物蛋白海绵供试品浸提液随浓度增加其吸光度（A值）增高，即NIH3T3细胞增殖作用增加，与空白组比较差异有统计学意义(P<0.01)，与bFGF标准品比较差异无统计学意义（P>0.05）；bFGF标准品和bFGF生物蛋白海绵在该方法中均存在量效关系，且符合四参数方程式：y=（A－D）∕［1+（X／C）B］+D；常温条件下储存2、3和4年生物蛋白海绵体与低温储存30 d比较，其生物学活性效价(U?mL-1)下降率分别为0.5%、0.6%和0.8%，均低于15%的国家标准。结论：bFGF生物蛋白海绵在体外能够促进NIH3T3细胞增殖，具有良好生物学活性；应用NIH3T3细胞检测bFGF生物蛋白海绵体外生物学活性可作为常规检测方法；bFGF生物蛋白海绵常温储存2~4年稳定性良好。

Abstract:

Abstract:Objective To establish the method for detection of biological activity of basic fibroblast growth factor(bFGF )protein sponge in vitro and investigate the stability of bFGF protein sponge stored at room temperature.Methods The bFGF biological protein sponge was fused into the dilution of the test solution,the extract was obtained after centrifugation,the bFGF standard samples were used as control,and the bFGF protein sponge was cultivated with NIH3T3 cells.MTT assay was used to dected the proflieration of NIH3T3 cells induced with bFGF biological protein sponge extract.The bFGF protein sponge stored for 2,3,4 years at room temperature was used as the test materials,the bFGF biological protein sponge stored for 30 d at low temperature as standard control,the culture medium without bFGF protein sponge and bFGF as blank control,the potency of biological activity of each group was detected.The stability of biological activities in vitro of bFGF protein sponge stored for different time were compared.Results Compared with blank control,the A value of bFGF biological protein sponge extract was increased with the increasing of concentration,in other words the NIH3T3 cell proliferation was increased(P<0.01);compared with bFGF standard control, the difference was not significant (P> 0.05),either bFGF standard or bFGF biological protein sponge showed a dose-response relationship by the developed method,and the determination result complied with the following four-parameter equation:y =（A-D）／［1 +（X ／ C）B］+D. Compared with protein sponge stored for 30 d at low temperature,the descending rate of biological activity titers (U?mL-1) of the protein sponge stored for 2 to 4 years at room temperature were 0.5%,0.6% and 0.8%,lower than the national standard of 15%. Conclusion The NIH3T3 cell proliferation could be applied for detection of the biological activity of bFGF protein sponge in vitro,it could be used as the routine detection method for bFGF protein sponge,the stabilities of bFGF protein sponge stored for 2-4 years at room temperature are good.

Key words: basic fibroblast growth factor, biologic protein sponge;biological activitydrug

中图分类号:

碱性成纤维细胞生长因子

焦悦|王晓杰|张睿|蔡敏倩|李婷婷|高丽昌|王珊珊|李海燕|李校堃. bFGF生物蛋白海绵体外生物学活性检测方法的建立及其稳定性评价[J]. J4, 2011, 37(1): 171-174.

JIAO Yue|WANG Xiao-jie|ZHANG Rui|CAI Min-qian|LI Ting-ting|GAO Li-cha|WANG Shan-. Establishment of method for detection of biological activity of |biological |protein sponge of |basic fibroblast growth factor |and stability evaluation[J]. J4, 2011, 37(1): 171-174.

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