High density cultivation of a recombinant CD-1 cell line producing prourokinase using a Biosilon microcarrier culture system.
CD-1, a genetically-engineered CHO cell line, was cultivated with a Biosilon microcarrier culture system. We successfully cultivated CD-1 cells to a very high density (over 1 x 10(7) cells/ml). Prourokinase was stably secreted at about 180 IU/10(6) cells/24 h. Experiments showed that CD-1 cells growing on Biosilon microcarriers were able to spontaneously release from the microcarriers, then reattach and proliferate on fresh microcarriers. This makes it very easy to scale up production. The microcarriers could be reused several times without affecting adhesion, proliferation and prourokinase secretion. With CM-PECC membrane radial flow chromatography and MPG chromatography, the prourokinase in conditioned medium could be purified to a specific activity of 1 x 10(5) IU/mg of protein. The purification factor was about 600 fold, and approximately 90% of the biological activity was recovered.
Xiao
C
C
Institute of Biotechnology, Academy of Military Medical Sciences, Beijing.
Huang
Z
Z
Zhang
Z
Z
Ye
J
J
Gao
L
L
Guo
Z
Z
Cheng
D
D
Zhou
H
H
Kong
W
W
D000818:Animals; D016466:CHO Cells / Q000378:metabolism*; D002452:Cell Count; D006224:Cricetinae; D003470:Culture Media; D004792:Enzyme Precursors / Q000378:metabolism; D005818:Genetic Engineering; D010957:Plasmids; D011137:Polystyrenes; D011994:Recombinant Proteins / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D003470:Culture Media; D004792:Enzyme Precursors; D011137:Polystyrenes; D011994:Recombinant Proteins; C024075:styrofoam; D014568:Urokinase-Type Plasminogen Activator; C067551:saruplase