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High yield antibody production in a disposable WAVE Bioreactor™
Process intensification using perfusion culture
Christian Kaisermayer1, Jianjun Yang2 1 GE Healthcare Europe GmbH, Vienna, Austria ([email protected]), 2 GE China Research and Development Center Co. Ltd. Shanghai First published in May 2011 at the 22nd ESACT meeting in Vienna, Austria
2 /High yield antibody production in a disposable WAVE Bioreactor
Introduction
• Perfusion processes provide high volumetric productivity by retaining a large cell number in a relatively small volume.
• These characteristics can be used for process intensification to obtain the same amount of product from a considerably smaller bioreactor compared to a standard batch or fedbatch process.
3 /High yield antibody production in a disposable WAVE Bioreactor
Introduction • Drosophila Schneider 2 (S2) insect cells producing a
recombinant antibody were cultivated in disposable WAVE Bioreactor™.
• Cell growth and protein production in a perfusion and a batch process were compared.
• Additionally the implication of both cultivation modes on the cost of consumables in the upstream process was evaluated.
4 /High yield antibody production in a disposable WAVE Bioreactor
Cell cultivation and reactor setups • S2 cells were transfected to produce a monoclonal antibody
against hemagglutinin of influenza H5N1.
• The cells were grown in commercial serum free medium. Inoculum was prepared in shake flasks.
• Cultivation was done in disposable WAVE Bioreactor™ at working volumes of 0.85 L.
• Oxygen was supplied by adjusting agitation and increasing O2 concentration in the headspace.
• In perfusion, cell retention was achieved by a filter integrated into the bioreactor.
• The perfusion rate was controlled via an integrated loadcell that allowed periodic addition and removal of equal amounts of feed and harvest.
5 /High yield antibody production in a disposable WAVE Bioreactor
Results
Fig. 1: Cell concentration and viability. Batch culture in red, perfusion culture in blue. Circles: cell concentration, triangles: viability. Bold line indicates perfusion rate.
6 /High yield antibody production in a disposable WAVE Bioreactor
Results • Perfusion had a drastic effect on cell concentration and
viability.
• As shown in Fig. 1, a tenfold higher cell concentration than in the batch culture was reached.
• Additionally a viability higher than 95% was maintained throughout the process.
• In combination this led to a 10 times larger viable cell integral for the perfusion culture.
7 /High yield antibody production in a disposable WAVE Bioreactor
Results
Fig. 2: Cell specific productivity (bars), specific perfusion rate (line) and IgG concentration (dots) during perfusion culture
8 /High yield antibody production in a disposable WAVE Bioreactor
Results • Utilization of cell culture medium was optimized by adjusting
the perfusion rate to ensure maximum product concentration but at the same time avoid nutrient limitation of the cells.
• The sudden decrease of the cell specific productivity on day 19 (Fig. 2) probably is the result of such a limit.
• A residual glucose concentration of about 3 g/L had to be maintained to avoid depletion of other nutrients.
• With the increased perfusion rate on day 20 and 21 cells recovered, reaching at least the same specific productivity as during the initial phase of the recombinant protein production.
9 /High yield antibody production in a disposable WAVE Bioreactor
Results
Fig. 3: IgG concentration (circles) and volumetric productivity (bar graph). Protein production was induced on day 10. Batch culture in red, closed symbols, perfusion culture in blue, open symbols
10 /High yield antibody production in a disposable WAVE Bioreactor
Results • Volumetric productivity (STY) was considerably higher in
perfusion culture as shown in Fig. 3.
• One of the reasons was the higher viable cell concentration, additionally also the cell specific productivity was improved more than twofold compared to the batch culture (Fig. 4).
• The high and stable productivity in perfusion culture can most likely be attributed to the consistent metabolite concentrations.
• As shown in Fig. 3 there were 6 days during the perfusion culture where a protein output of more than 1 g per liter working volume was achieved.
11 /High yield antibody production in a disposable WAVE Bioreactor
Results
Fig. 4: Comparison of cell concentration, productivity and consumable cost in both culture modes
12 /High yield antibody production in a disposable WAVE Bioreactor
Conclusions
• Perfusion allowed a 20-fold increase of the volumetric productivity compared to batch culture
• The upstream consumable cost per 100 mg IgG decreased by 85%
• A viable cell concentration of about 1*108 c/ml was maintained for one week
• Use of a disposable bioreactor allows short turnover times, improving equipment utilization
• Despite the high cell concentration, no DO limits or clogging of the retention filter were observed during 3 weeks process time
13 /High yield antibody production in a disposable WAVE Bioreactor
Acknowledgments The S 2 cell line used in this study, was generously provided by Lulan Wang and Paul Zhou, Institut Pasteur, Shanghai
© 2011 General Electric Company - All rights reserved.
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WAVE Bioreactor is a trademark of GE Healthcare companies.
All goods and services are sold subjects to the terms and conditions of sale of the company within GE Healthcare which supplies them.
A copy of these terms and conditions is available on request.
Contact your local GE Healthcare representative for the most current information.
First published May 2011.