[Production of u-PA with rCHO cell culture on porous microcarriers in serum-free growth medium].
A novel technique was developed to deal with apoptosis in large-scale animal cell culture. By means of replacing part of Cytopore porous microcarriers at regular intervals, a rCHO cell line, which produces urokinase-type plasminogen activitor(u-PA), was cultivated continuously with serum-free medium in a 30 L stirred tank for 91 days. The cell density was maintained at (1.3-2.6) x 10(7)/mL, and > 90% of cells was viable. In order to reduce the effect of cell density on cell growth and expression, a cyclic pressure oscillation was exerted on a 7.5 L reactor headspace to enhance cell expression at high cell density to a certain extent. During the 67 days of medium-replacement culture, the maximal cell density reached 2.64 x 10(7)/mL, and cell viability was always kept above 95% when combined with microcarrier-replacement. Compare to control culture, culture with cyclic pressure oscillation could enhance cell expression level and reduce the ratio of glucose metabolized anaerobically to produce lactate. With four-step purification process, about 80 g u-PA(approximately 90% scu-PA) was recovered from approximately 2100 liters supernatant which contained approximately 135 g u-PA.
Hu
X W
XW
Institute of Biotechnology, Academy of Military Medical Sciences, Beijing.
Xiao
C Z
CZ
Li
Z H
ZH
Guo
Z X
ZX
Gao
L H
LH
Zhang
Z G
ZG
Xu
Z P
ZP
Wang
F
F
D000818:Animals; D016466:CHO Cells; D002455:Cell Division; D006224:Cricetinae; D016895:Culture Media, Serum-Free; D014568:Urokinase-Type Plasminogen Activator / Q000096:biosynthesis*
D016895:Culture Media, Serum-Free; D014568:Urokinase-Type Plasminogen Activator