Optimization of Culture Conditions for MDCK Cell Micro-carriers

doi:10.13560/j.cnki.biotech.bull.1985.2018-0491

Abstract

Abstract: Studying the effect of cell inoculation quantity,stirring speed and microcarrier concentration on the MDCK cells in micro-carrier culture and optimizing the conditions of culturing MDCK cells micro-carrier during the age of the largest proliferation is of great significance for the separation of vaccine from virus,aiming at increasing the efficiency in separating vaccine from viruses. In this study,MDCK cells were cultured with a micro-vector and cultured at different stirring speed,concentration of the micro-vector,and amount of cell inoculation,and the optimal culture conditions were determined by sampling and counting every 24 h. The results showed that MDCK cells proliferation was faster and cell density was larger up to 16.5 x 10⁶cells/mL while cell inoculation in 20 balls,stirring speed in 45 r/min,and carrier concentration in 2 g/L,which is suitable for MDCK cells proliferation and growth.

Key words: MDCK cells, cell volume, micro-carrier culture

MA Qi-cai, WU Wen-li, MA Fu-long, PAN He-ping. Optimization of Culture Conditions for MDCK Cell Micro-carriers[J]. Biotechnology Bulletin, 2018, 34(12): 90-95.

References

[1] Madin SH, Darby NB Jr.Established kidney cell lines of normal adult bovine and ovine origin[J]. Proc Soc Exp Biol Med, 1958, 98(3):574-576.
[2] 过琴媛, 王辉, 沈心亮. 微载体培养动物细胞技术的研究进展[J]. 微生物学免疫学进展, 2007, 35(1):73-75.
[3] 黄锭, 赵亮, 谭文松. 犬肾细胞MDCK无血清贴壁及单细胞悬浮培养[J]. 生物工程学报, 2011, 27(4):645-652.
[4] 贾涵婧, 王逸群, 黄锭, 等. 微载体浓度与细胞接种密度对MDCK细胞生长的影响[J]. 中国生物制品学杂志, 2014, 27(9):1138-1144.
[5] Trabelsi K, Majoul S, Rourou S, et al Development of a measles vaccine production process in MRC-5 cells grown on Cytodex1 microcarriers and in a stirred bioreactor[J]. Applied Microbiology & Biotechnology, 2012, 93(3):1031-1040.
[6] Rourou S, Ark AVD, Majoul S, et al.A novel animal-component-free medium for rabies virus production in Vero cells grown on Cytodex 1 microcarriers in a stirred bioreactor[J]. Applied Microbiology & Biotechnology, 2009, 85(1):53.
[7] Wu SC, Liu CC, Lian WC.Optimization of microcarrier cell culture process for the inactivated enterovirus type 71 vaccine development[J]. Vaccine, 2004, 22(29-30):3858-3864.
[8] 龚迪, 易小萍, 张元兴. MDCK细胞微载体悬浮培养放大工艺研究[J]. 中国生物工程杂志, 2012, 32(9):55-60.
[9] Arifin MA, Mel M, Abdul Karim MI, et al.Production of Newcastle disease virus by Vero cells grown on cytodex1 microcarriers in a 2-litre stirred tank bioreactor[J]. Journal of Biomedicine & Biotechnology, 2015, 2010(2010):586363.
[10] 林美玲. Vero细胞低血清微载体悬浮培养模式的优化及溶瘤单纯疱疹病毒生产工艺研究[D]. 上海:华东理工大学, 2014.
[11] 蒋春雷, 王笑利. 介绍一种少量冻存细胞复苏技术[J]. 生物学杂志, 1991(5):27-27.
[12] 俞索静, 吴承亮, 金红婷, 等. 破骨细胞的传代、冻存与复苏[J]. 中国骨伤, 2017, 30(5):463-469.
[13] 黄锭, 赵亮, 谭文松. 犬肾细胞MDCK无血清贴壁及单细胞悬浮培养[J]. 生物工程学报, 2011, 27(24):645-652.
[14] 黄锭. 用于流感疫苗生产的MDCK细胞无血清单细胞悬浮培养体系的开发[D]. 上海:华东理工大学, 2010.
[15] 古少鹏, 赵素芬. 王运盛等鸡胚盲肠上皮细胞的传代培养[J]. 畜牧兽医学报, 2011, 42(3):409-415.
[16] 李吉霞. 小鼠胚胎生殖细胞的分离与培养[D]. 杨凌:西北农林科技大学, 2005.
[17] 曹锴. 草鱼脂肪细胞原代及传代培养[D]. 重庆:重庆师范大学, 2011.
[18] 张志侠. 如何让学生学会使用血球计数板[J]. 科学大众:科学教育, 2011(7):5-5.
[19] 邬扬镰, 胡文炳. 血球计数板的划线及深度测量[J]. 临床检验杂志, 1985(2):34.
[20] Clark J M, Hirtenstein MD.Optimizing culture conditioned for the production of animal cells in microcarrier culture[J]. Annals of the New York Academy of Sciences, 1981, 369(1):33-46.
[21] 黄锭, 刘旭平, 范里, 等. 感染时间对MDCK细胞中H1N1甲型流感病毒扩增过程的影响[J]. 生物技术通报, 2015, 31(11):243-250.
[22] 刘鹏, 李佳林, 马超, 等. H1N1流感病毒在微载体培养MDCK细胞上增殖的研究[J]. 微生物学免疫学进展, 2013, 41(1):12-15.
[23] 陈以衡, 陶姝宇, 刘旭平, 谭文松. 微载体浓度与细胞接种密度对ST细胞生长的影响[J]. 生物技术通报, 2016, 32(4):242-250.
[24] 黄锭, 赵亮, 谭文松. 犬肾细胞MDCK无血清贴壁及单细胞悬浮培养[J]. 生物工程学报, 2011, 27(4):645-652.