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Animal Cell Biotechnology pp 125–140Cite as

Development of Mammalian Cell Perfusion Cultures at Lab Scale: From Orbitally Shaken Tubes to Benchtop Bioreactors

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 2095))

Abstract

This chapter introduces the necessary concepts to develop mammalian cell perfusion cultures for the expression of therapeutic proteins at lab scale. We highlight the operation of the orbitally shaken tubes and of a classical glass vessel reactor system coupled to an external alternating tangential flow (ATF) device. Two different experiments can be performed in the shake-tube system: (1) the VCDmax experiment exploring the maximum achievable viable cell density at a given medium exchange rate and (2) the VCDSS experiment for the prediction of process performance at constant viable cell density and a given medium exchange rate for the design of the benchtop bioreactor process. In addition, the operation of the benchtop system is discussed containing start-up and control procedures for a long-term production run.

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References

  1. Lute S, Agarabi C, Johnson S, Chavez B, Brorson K (2017) FDA/OBP laboratory research to support continuous bioprocessing.In:Integrated Continuous Biomanufacturing III. Suzanne Farid, University College London, United Kingdom Chetan Goudar, Amgen, USA Paula Alves, IBET, Portugal Veena Warikoo, Axcella Health, Inc., USA Eds, ECI Symposium Series

    Google Scholar 

  2. Zydney AL (2015) Perspectives on integrated continuous bioprocessing – opportunities and challenges. Curr Opin Chem Eng 10:8–13. https://doi.org/10.1016/j.coche.2015.07.005

    Article  Google Scholar 

  3. Konstantinov KB, Cooney CL (2015) White paper on continuous bioprocessing. May 20–21, 2014 continuous manufacturing symposium. J Pharm Sci 104:813–820. https://doi.org/10.1002/jps.24268

    Article  CAS  PubMed  Google Scholar 

  4. Croughan MS, Konstantinov KB, Cooney C (2015) The future of industrial bioprocessing: batch or continuous? Biotechnol Bioeng 112:648–651. https://doi.org/10.1002/bit.25529

    Article  CAS  PubMed  Google Scholar 

  5. Subramanian G (2018) Continuous biomanufacturing: innovative technologies and methods. Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

    Google Scholar 

  6. Farid SS, Pollock J, Ho SV (2014) Evaluating the economic and operational feasibility of continuous processes for monoclonal antibodies. In: Continuous processing in pharmaceutical manufacturing. Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim, pp 433–456

    Google Scholar 

  7. Steinebach F, Ulmer N, Wolf M, Decker L, Schneider V, Wälchli R, Karst D, Souquet J, Morbidelli M (2017) Design and operation of a continuous integrated monoclonal antibody production process. Biotechnol Prog 33:1303–1313. https://doi.org/10.1002/btpr.2522

    Article  CAS  PubMed  Google Scholar 

  8. Sawyer D, Sanderson K, Lu R, Daszkowski T, Clark E, Mcduff P, Astrom J, Heffernan C, Duffy L, Poole S, Ryll T, Sheehy P, Strachan D, Souquet J, Beattie D, Pollard D, Stauch O, Bezy P, Sauer T, Boettcher L, Simpson C, Dakin J, Pitt S, Boyle A (2017) Biomanufacturing technology roadmap – overview

    Google Scholar 

  9. Walther J, Godawat R, Hwang C, Abe Y, Sinclair A, Konstantinov K (2015) The business impact of an integrated continuous biomanufacturing platform for recombinant protein production. J Biotechnol 213:3–12. https://doi.org/10.1016/j.jbiotec.2015.05.010

    Article  CAS  PubMed  Google Scholar 

  10. Xenopoulos A (2015) A new, integrated, continuous purification process template for monoclonal antibodies: process modeling and cost of goods studies. J Biotechnol 213:42–53. https://doi.org/10.1016/j.jbiotec.2015.04.020

    Article  CAS  PubMed  Google Scholar 

  11. Xu S, Gavin J, Jiang R, Chen H (2017) Bioreactor productivity and media cost comparison for different intensified cell culture processes. Biotechnol Prog 33:867–878. https://doi.org/10.1002/btpr.2415

    Article  CAS  PubMed  Google Scholar 

  12. Reay D, Ramshaw C, Harvey A (2013) Process intensification: engineering for efficiency, sustainability and flexibility. Butterworth-Heinemann, Oxford

    Book  Google Scholar 

  13. Moulijn JA, Stankiewicz A, Grievink J, Górak A (2008) Process intensification and process systems engineering: a friendly symbiosis. Comput Chem Eng 32:3–11. https://doi.org/10.1016/j.compchemeng.2007.05.014

    Article  CAS  Google Scholar 

  14. Bödeker BGD, Newcomb R, Yuan P, Braufman A, Kelsey W (1994) Production of recombinant factor VIII from perfusion cultures: i. Large-scale fermentation. In: Spier RE, Griffiths JB, Berthold W (eds) Animal cell technology. Butterworth-Heinemann, Oxford, pp 580–583

    Chapter  Google Scholar 

  15. Langer ES (2011) Trends in perfusion bioreactors. Bioprocess Int 9:10

    Google Scholar 

  16. Bonham-Carter J, Shevitz J et al (2011) A brief history of perfusion biomanufacturing. BioProcess Int 9:24–30

    Google Scholar 

  17. Barrett S, Chang A, Bandow N (2017) Intensification of a multi-product perfusion platform through medium and process development. In: Farid S, Goudar C, Alves P, Warikoo P (eds) Integrated continuous biomanufacturing III. Engineering Conferences International, Cascais, Portugal

    Google Scholar 

  18. Deschênes J-S, Desbiens A, Perrier M, Kamen A (2006) Use of cell bleed in a high cell density perfusion culture and multivariable control of biomass and metabolite concentrations. Asia-Pacific. J Chem Eng 1:82–91. https://doi.org/10.1002/apj.10

    Article  Google Scholar 

  19. Lin H, Leighty RW, Godfrey S, Wang SB (2017) Principles and approach to developing mammalian cell culture media for high cell density perfusion process leveraging established fed-batch media. Biotechnol Prog 33:891–901. https://doi.org/10.1002/btpr.2472

    Article  CAS  PubMed  Google Scholar 

  20. Wolf MKF, Closet A, Bzowska M, Bielser J-M, Souquet J, Broly H, Morbidelli M (2018) Improved performance in mammalian cell perfusion cultures by growth inhibition. Biotechnol J 14(2):e1700722. https://doi.org/10.1002/biot.201700722

    Article  CAS  PubMed  Google Scholar 

  21. Konstantinov K, Goudar C, Ng M, Meneses R, Thrift J, Chuppa S, Matanguihan C, Michaels J, Naveh D (2006) The “push-to-low” approach for optimization of high-density perfusion cultures of animal cells. Adv Biochem Eng Biotechnol 101:75–98. https://doi.org/10.1007/10_016

    Article  CAS  PubMed  Google Scholar 

  22. Bausch M, Schultheiss C, Sieck JB (2018) Recommendations for comparison of productivity between fed-batch and perfusion processes. Biotechnol J 14(2):e1700721. https://doi.org/10.1002/biot.201700721

    Article  CAS  PubMed  Google Scholar 

  23. Dowd JE, Jubb A, Kwok KE, Piret JM (2003) Optimization and control of perfusion cultures using a viable cell probe and cell specific perfusion rates. Cytotechnology 42:35–45. https://doi.org/10.1023/A:1026192228471

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Xu S, Chen H (2016) High-density mammalian cell cultures in stirred-tank bioreactor without external pH control. J Biotechnol 231:149–159. https://doi.org/10.1016/j.jbiotec.2016.06.019

    Article  CAS  PubMed  Google Scholar 

  25. Karst DJ, Serra E, Villiger TK, Soos M, Morbidelli M (2016) Characterization and comparison of ATF and TFF in stirred bioreactors for continuous mammalian cell culture processes. Biochem Eng J 110:17–26. https://doi.org/10.1016/j.bej.2016.02.003

    Article  CAS  Google Scholar 

  26. Goletz S, Stahn R, Kreye S (2016) Small scale cultivation method for suspension cells

    Google Scholar 

  27. Gomez N, Ambhaikar M, Zhang L, Huang C-J, Barkhordarian H, Lull J, Gutierrez C (2017) Analysis of Tubespins as a suitable scale-down model of bioreactors for high cell density CHO cell culture. Biotechnol Prog 33:490–499. https://doi.org/10.1002/btpr.2418

    Article  CAS  PubMed  Google Scholar 

  28. Villiger-Oberbek A, Yang Y, Zhou W, Yang J (2015) Development and application of a high-throughput platform for perfusion-based cell culture processes. J Biotechnol 212:21–29. https://doi.org/10.1016/j.jbiotec.2015.06.428

    Article  CAS  PubMed  Google Scholar 

  29. Wolf MKF, Lorenz V, Karst DJ, Souquet J, Broly H, Morbidelli M (2018) Development of a shake tube-based scale-down model for perfusion cultures. Biotechnol Bioeng 115:2703–2713. https://doi.org/10.1002/bit.26804

    Article  CAS  PubMed  Google Scholar 

  30. Sieck JB, Schild C, von Hagen J (2017) Perfusion formats and their specific medium requirements. In: Continuous biomanufacturing – innovative technologies and methods, pp 171–200. https://doi.org/10.1002/9783527699902.ch7

    Chapter  Google Scholar 

  31. Chotteau V (2017) Process development in screening scale bioreactors and perspectives for very high cell density perfusion. In: Integrated Continuous Biomanufacturing III. Suzanne Farid, University College London, United Kingdom Chetan Goudar, Amgen, USA Paula Alves, IBET, Portugal Veena Warikoo, Axcella Health, Inc., USA Eds, ECI Symposium Series

    Google Scholar 

  32. Zoro B, Tait A, Carpio M, McHugh K (2018) Development of a novel automated perfusion mini-bioreactor ambr® 250 perfusion. In: A. Robinson, PhD, Tulane University R. Venkat, PhD, MedImmune E. Schaefer, ScD, J&J Janssen,editors.Cell Culture Engineering XVI, ECI Symposium Series, 2018

    Google Scholar 

  33. Villiger TK, Neunstoecklin B, Karst DJ, Lucas E, Stettler M, Broly H, Morbidelli M, Soos M (2018) Experimental and CFD physical characterization of animal cell bioreactors: from micro- to production scale. Biochem Eng J 131:84–94. https://doi.org/10.1016/j.bej.2017.12.004

    Article  CAS  Google Scholar 

  34. Garcia-Ochoa F, Gomez E (2009) Bioreactor scale-up and oxygen transfer rate in microbial processes: an overview. Biotechnol Adv 27:153–176. https://doi.org/10.1016/j.biotechadv.2008.10.006

    Article  CAS  PubMed  Google Scholar 

  35. Russell TWF, Robinson AS, Wagner NJ (2008) Mass and heat transfer: analysis of mass contactors and heat exchangers. Cambridge University Press, New York

    Book  Google Scholar 

  36. Vogg S, Wolf MKF, Morbidelli M (2018) Continuous and integrated expression and purification of recombinant antibodies. In: Hacker DL (ed) Methods in molecular biology. Springer, New York, NY, pp 147–178

    Google Scholar 

  37. Clincke M, Mölleryd C, Zhang Y, Lindskog E, Walsh K, Chotteau V (2013) Very high density of CHO cells in perfusion by ATF or TFF in WAVE bioreactorTM. Part I. Effect of the cell density on the process. Biotechnol Prog 29:754–767. https://doi.org/10.1002/btpr.1704

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Fan Y, Ley D, Andersen MR (2018) Fed-Batch CHO Cell Culture for Lab-Scale Antibody Production. Methods Mol Biol 1674:147–161. https://doi.org/10.1007/978-1-4939-7312-5

    Article  CAS  PubMed  Google Scholar 

  39. Zhu L, Song B, Wang Z, Monteil DT, Shen X, Hacker DL, De Jesus M, Wurm FM (2017) Studies on fluid dynamics of the flow field and gas transfer in orbitally shaken tubes. Biotechnol Prog 33:192–200. https://doi.org/10.1002/btpr.2375

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work has been supported by the KTI (CTI) Program of the Swiss Economic Ministry (Project 19190.2 PFIW-IW). The authors declare that they have no conflicts of interest pertaining to the contents of this article.

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Authors and Affiliations

  1. Institute for Chemical and Bioengineering, ETH Zurich, Zurich, Switzerland

    Moritz Wolf & Massimo Morbidelli

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  1. Moritz Wolf
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  2. Massimo Morbidelli
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Correspondence to Moritz Wolf .

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Editors and Affiliations

  1. Institute of Bioprocess and Biosystems Engineering, Hamburg University of Technology (TUHH), Hamburg, Germany

    Ralf Pörtner

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Wolf, M., Morbidelli, M. (2020). Development of Mammalian Cell Perfusion Cultures at Lab Scale: From Orbitally Shaken Tubes to Benchtop Bioreactors. In: Pörtner, R. (eds) Animal Cell Biotechnology. Methods in Molecular Biology, vol 2095. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0191-4_8

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  • DOI: https://doi.org/10.1007/978-1-0716-0191-4_8

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0190-7

  • Online ISBN: 978-1-0716-0191-4

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