Materials and Methods
Overview of setting-up procedures
Day -6: Inoculum production with CHO suspension cells (CHO XM111-10) in shake flasks.
Day -1/0: Bioreactor and medium preparation.
Day 0: Inoculation and starting-up of the glass bioreactor with seeding density of 0.6 x 106 cells/mL-1 in 1.0 L ChoMaster® HP-1 growth medium (that was supplemented with 2.0 g·L-1 Pluronic F-68 and 2.5 mg L-1 tetracycline).
Day 2: Sampling, addition of 1.0 L ChoMaster HP-5 growth medium. Increase of impeller speed to maintain constant specific power inputs.
Day 3: Sampling, sedimentation of cells and removal of supernatant for medium exchange to tetracycline-free ChoMaster HP-5 production medium.
Day 4: Sampling, temperature shift from 37°C to 31°C.
Days 5–9: Sampling, analytics, and harvest.
For the seed inoculum production and the start of the cultivation in the glass bioreactor, ChoMaster HP-1 medium that was supplemented with 2.0 gL-1 Pluronic F-68 and 2.5 mgL-1 tetracycline (Cell Culture Technologies) was used. The feeding was realized with ChoMaster HP-5 growth medium, and the SEAP secretion was induced by medium exchange to tetracycline-free ChoMaster HP-5 production medium.