- Poster presentation
- Open Access
Highly efficient inoculum propagation in perfusion culture using WAVE Bioreactor™ systems
© Kaisermayer et al.; licensee BioMed Central Ltd. 2013
- Published: 4 December 2013
- Batch Culture
- Split Ratio
- Shorten Process Time
- Perfusion Culture
- High Cell Concentration
A perfusion-based process was developed to increase the split ratio during the scale-up of CHO-S™ cell cultures. Fedbatch cultures were inoculated with cells propagated in either batch or perfusion cultures. All cultures were grown in disposable Cellbag™ bioreactors using the WAVE Bioreactor system. Cell concentrations of 4.8 × 107 cells/mL were achieved in the perfusion culture, whereas the final cell concentration in the batch culture was 5.1 × 106 cells/mL. The higher cell concentration of the perfusion culture allowed for a more than six-fold increase of the split ratio to about 1:30. The method described here, can reduce the number of required expansion steps and eliminate the need for one or two bioreactors in the seed train. Single-use bioreactors at benchtop scale can be used for direct inoculation of production bioreactors. Alternatively, high biomass concentrations accumulated in perfusion culture can be used to seed production vessels at increased cell concentrations. Thus, the process time in these bioreactors, which often is the bottleneck in plant throughput, can be shortened.
CHO-S cells (Life Technlologies)
Cultivation medium and feed concentrate: T13 and T13-F (Shanghai Hankang Biotech Co.)
WAVE Biorereactor 20/50 system (GE Healthcare)
Cellbag bioreactors (GE Healthcare)
Batch and fed-batch cultivations were run in Cellbag 10 L bioreactors, perfusion cultures in Cellbag 2 L bioreactors. Cultivation conditions: T 37°C, pH 7.10, DO > 40%, agitation for all cultures 25 rpm/6°.
Analytics: Cell concentration and viability, glucose and lactate concentration. Perfusion and feed rates were adjusted to maintain the residual glucose concentration above 0.5 g/L.
Comparison of fed-batch cultures
FB seeded from batch
Cell conc. at cell removal [c/mL]
2.2 × 106
2.3 × 107
Inoculum conc. [c/mL]
4.1 × 105
7.4 × 105
Process time [d]
Peak cell conc. [c/mL]
1.4 × 107
1.7 × 107
Av. μ during growth phase [d-1]
Perfusion culture maintained cells in exponential growth phase for an extended period of time compared with batch culture.
The high cell concentrations obtained in perfusion culture can substantially increase the split ratio, thus, minimizing the number of vessels needed in the seed train.
Alternatively, the production bioreactor can be inoculated at high cell concentration, which can help shortening process time in the production vessel and improving facility utilization.
One WAVE Bioreactor 20/50 system, run in perfusion mode at the maximum operating volume of 25 L, could provide inoculum for a 2000 L bioreactor.
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