Studies On The Production Of Pro-UK Using Insect Cell Culturein Bioreactor

A new production technology of prourokinase（pro-UK） by insect cell culture and baculovirus was established using high density cells attachment culture of sf9 insect cells in a celligen-packed bed basket bioreactor and fibracel disks lamellar carriers. In the research, the effect of carriers treatment on sf9 cells attachment was studied,then the method that carriers has to be soaked in serum-supplemented medium of TNM-FN over 2 hours was determined. 3.0～5.0×10⁵ cells/ml cell density was applied as seeded concentration of insect cell culture in bioreactors, and stirring speed was controlled between 30 and 50rpm for 6 hours after seeding. It was proved that insect cell culture don’t need CO₂, cell growth will be inhibited intensively by CO₂,. This paper also compared the infection stage and dosage of baculovirus AcMNPV, then cultivation for 4～5 days was chosen as optimal stage of infection, and the multiplicity of infection（MOI） is at among 1～2MOI. Perfusion culture was prove to be better than batch culture. 1.5×10⁷cells/ml cell density was reached in perfusion culture, and pro-UK could reach as high as 1432IU/ml when the harvest rate is at 0.5V/day, and pro-UK expressed often lasts more than 500 IU/ml for a long time （8～10days）. This study lays a technology foundation for the large-scale production of pro-UK.