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Turbidimetry and Dielectric Spectroscopy as Process Analytical Technologies for Mammalian and Insect Cell Cultures

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Animal Cell Biotechnology

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

Abstract

The production of biopharmaceuticals in cell culture involves stringent controls to ensure product safety and quality. To meet these requirements, quality by design principles must be applied during the development of cell culture processes so that quality is built into the product by understanding the manufacturing process. One key aspect is process analytical technology, in which comprehensive online monitoring is used to identify and control critical process parameters that affect critical quality attributes such as the product titer and purity. The application of industry-ready technologies such as turbidimetry and dielectric spectroscopy provides a deeper understanding of biological processes within the bioreactor and allows the physiological status of the cells to be monitored on a continuous basis. This in turn enables selective and targeted process controls to respond in an appropriate manner to process disturbances. This chapter outlines the principles of online dielectric spectroscopy and turbidimetry for the measurement of optical density as applied to mammalian and insect cells cultivated in stirred-tank bioreactors either in suspension or as adherent cells on microcarriers.

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References

  1. International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (2009) ICH Q8(R2): pharmaceutical development. https://www.ich.org/fileadmin/Public_Web_Site/ICH_Products/Guidelines/Quality/Q8_R1/Step4/Q8_R2_Guideline.pdf

  2. International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (1999) ICH Q6A: specifications: test procedures and acceptance criteria for new drug substances and new drug products: chemical substances. https://www.ich.org/fileadmin/Public_Web_Site/ICH_Products/Guidelines/Quality/Q6A/Step4/Q6Astep4.pdf

  3. Beutel S, Henkel S (2011) In situ sensor techniques in modern bioprocess monitoring. Appl Microbiol Biotechnol 91(6):1493–1505. https://doi.org/10.1007/s00253-011-3470-5

    Article  CAS  Google Scholar 

  4. Biechele P, Busse C, Solle D et al (2015) Sensor systems for bioprocess monitoring. Eng Life Sci 15(5):469–488. https://doi.org/10.1002/elsc.201500014

    Article  CAS  Google Scholar 

  5. Eibl R, Eibl D, Pörtner R et al (eds) (2009) Cell and tissue reaction engineering. Principles and practice. Springer, Berlin, Heidelberg

    Google Scholar 

  6. Carvell JP, Dowd JE (2006) On-line measurements and control of viable cell density in cell culture manufacturing processes using radio-frequency impedance. Cytotechnology 50(1-3):35–48. https://doi.org/10.1007/s10616-005-3974-x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Kiviharju K, Salonen K, Moilanen U et al (2008) Biomass measurement online: the performance of in situ measurements and software sensors. J Ind Microbiol Biotechnol 35(7):657–665. https://doi.org/10.1007/s10295-008-0346-5

    Article  CAS  PubMed  Google Scholar 

  8. Justice C, Brix A, Freimark D et al (2011) Process control in cell culture technology using dielectric spectroscopy. Biotechnol Adv 29(4):391–401. https://doi.org/10.1016/j.biotechadv.2011.03.002

    Article  CAS  PubMed  Google Scholar 

  9. Zitzmann J, Weidner T, Eichner G et al (2018) Dielectric spectroscopy and optical density measurement for the online monitoring and control of recombinant protein production in stably transformed Drosophila melanogaster S2 cells. Sensors (Basel) 18:3. https://doi.org/10.3390/s18030900

    Article  CAS  Google Scholar 

  10. Wu P, Ozturk SS, Blackie JD et al (1995) Evaluation and applications of optical cell density probes in mammalian cell bioreactors. Biotechnol Bioeng 45(6):495–502. https://doi.org/10.1002/bit.260450606

    Article  CAS  PubMed  Google Scholar 

  11. Cervera AE, Petersen N, Lantz AE et al (2009) Application of near-infrared spectroscopy for monitoring and control of cell culture and fermentation. Biotechnol Prog 25(6):1561–1581. https://doi.org/10.1002/btpr.280

    Article  CAS  PubMed  Google Scholar 

  12. Qiu J, Arnold MA, Murhammer DW (2014) On-line near infrared bioreactor monitoring of cell density and concentrations of glucose and lactate during insect cell cultivation. J Biotechnol 173:106–111. https://doi.org/10.1016/j.jbiotec.2014.01.009

    Article  CAS  PubMed  Google Scholar 

  13. Marose S, Lindemann C, Scheper T (1998) Two-dimensional fluorescence spectroscopy: a new tool for on-line bioprocess monitoring. Biotechnol Prog 14(1):63–74. https://doi.org/10.1021/bp970124o

    Article  CAS  PubMed  Google Scholar 

  14. Abu-Absi NR, Kenty BM, Cuellar ME et al (2011) Real time monitoring of multiple parameters in mammalian cell culture bioreactors using an in-line Raman spectroscopy probe. Biotechnol Bioeng 108(5):1215–1221. https://doi.org/10.1002/bit.23023

    Article  CAS  PubMed  Google Scholar 

  15. Joeris K, Frerichs J-G, Konstantinov K et al (2002) In-situ microscopy: online process monitoring of mammalian cell cultures. Cytotechnology 38(1-3):129–134. https://doi.org/10.1023/A:1021170502775

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Bluma A, Höpfner T, Lindner P et al (2010) In-situ imaging sensors for bioprocess monitoring: state of the art. Anal Bioanal Chem 398(6):2429–2438. https://doi.org/10.1007/s00216-010-4181-y

    Article  CAS  PubMed  Google Scholar 

  17. Whelan J, Murphy E, Pearson A et al (2012) Use of focused beam reflectance measurement (FBRM) for monitoring changes in biomass concentration. Bioprocess Biosyst Eng 35(6):963–975. https://doi.org/10.1007/s00449-012-0681-9

    Article  CAS  PubMed  Google Scholar 

  18. Cole H, Demont A, Marison I (2015) The application of dielectric spectroscopy and biocalorimetry for the monitoring of biomass in immobilized mammalian cell cultures. Processes 3(2):384–405. https://doi.org/10.3390/pr3020384

    Article  CAS  Google Scholar 

  19. Aehle M, Kuprijanov A, Schaepe S et al (2011) Simplified off-gas analyses in animal cell cultures for process monitoring and control purposes. Biotechnol Lett 33(11):2103–2110. https://doi.org/10.1007/s10529-011-0686-5

    Article  CAS  PubMed  Google Scholar 

  20. Chen LZ, Nguang SK, Li XM et al (2004) Soft sensors for on-line biomass measurements. Bioprocess Biosyst Eng 26(3):191–195. https://doi.org/10.1007/s00449-004-0350-8

    Article  CAS  PubMed  Google Scholar 

  21. Ducommun P, Kadouri A, von Stockar U et al (2002) On-line determination of animal cell concentration in two industrial high-density culture processes by dielectric spectroscopy. Biotechnol Bioeng 77(3):316–323. https://doi.org/10.1002/bit.1197

    Article  CAS  PubMed  Google Scholar 

  22. Konstantinov K, Chuppa S, Sajan E et al (1994) Real-time biomass-concentration monitoring in animal-cell cultures. Trends in Biotechnology 12(8):324–333. https://doi.org/10.1016/0167-7799(94)90049-3

    Article  CAS  PubMed  Google Scholar 

  23. Junker BH, Reddy J, Gbewonyo K et al (1994) On-line and in-situ monitoring technology for cell density measurement in microbial and animal cell cultures. Bioprocess Eng 10(5-6):195–207. https://doi.org/10.1007/BF00369530

    Article  Google Scholar 

  24. Myers JA, Curtis BS, Curtis WR (2013) Improving accuracy of cell and chromophore concentration measurements using optical density. BMC Biophys 6(1):4. https://doi.org/10.1186/2046-1682-6-4

    Article  PubMed  PubMed Central  Google Scholar 

  25. Zhao L, Fu H-Y, Zhou W et al (2015) Advances in process monitoring tools for cell culture bioprocesses. Eng Life Sci 15(5):459–468. https://doi.org/10.1002/elsc.201500006

    Article  CAS  Google Scholar 

  26. Yardley JE, Kell DB, Barrett J et al (2000) On-line, real-time measurements of cellular biomass using dielectric spectroscopy. Biotechnol Genet Eng Rev 17(1):3–36. https://doi.org/10.1080/02648725.2000.10647986

    Article  CAS  PubMed  Google Scholar 

  27. Opel CF, Li J, Amanullah A (2010) Quantitative modeling of viable cell density, cell size, intracellular conductivity, and membrane capacitance in batch and fed-batch CHO processes using dielectric spectroscopy. Biotechnol Prog 26(4):1187–1199. https://doi.org/10.1002/btpr.425

    Article  CAS  PubMed  Google Scholar 

  28. Braasch K, Nikolic-Jaric M, Cabel T et al (2013) The changing dielectric properties of CHO cells can be used to determine early apoptotic events in a bioprocess. Biotechnol Bioeng 110(11):2902–2914. https://doi.org/10.1002/bit.24976

    Article  CAS  PubMed  Google Scholar 

  29. Schwan HP (1957) Electrical properties of tissue and cell suspensions, vol 5. Elsevier, Amsterdam, pp 147–209

    Google Scholar 

  30. Asami K (2002) Characterization of heterogeneous systems by dielectric spectroscopy. Progr Polym Sci 27(8):1617–1659. https://doi.org/10.1016/S0079-6700(02)00015-1

    Article  CAS  Google Scholar 

  31. Markx GH, Davey CL (1999) The dielectric properties of biological cells at radiofrequencies: applications in biotechnology. Enzyme Microbial Technol 25(3-5):161–171. https://doi.org/10.1016/S0141-0229(99)00008-3

    Article  CAS  Google Scholar 

  32. Ansorge S, Esteban G, Schmid G (2007) On-line monitoring of infected Sf-9 insect cell cultures by scanning permittivity measurements and comparison with off-line biovolume measurements. Cytotechnology 55(2-3):115–124. https://doi.org/10.1007/s10616-007-9093-0

    Article  PubMed  PubMed Central  Google Scholar 

  33. Zitzmann J, Sprick G, Weidner T et al (2017) Process optimization for recombinant protein expression in insect cells. In: Gowder SJT (ed) New insights into cell culture technology. InTech Open, London

    Google Scholar 

  34. Kost TA, Condreay JP, Jarvis DL (2005) Baculovirus as versatile vectors for protein expression in insect and mammalian cells. Nat Biotechnol 23(5):567–575. https://doi.org/10.1038/nbt1095

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Negrete A, Esteban G, Kotin RM (2007) Process optimization of large-scale production of recombinant adeno-associated vectors using dielectric spectroscopy. Appl Microbiol Biotechnol 76(4):761–772. https://doi.org/10.1007/s00253-007-1030-9

    Article  CAS  PubMed  Google Scholar 

  36. Zeiser A, Elias CB, Voyer R et al (2000) On-line monitoring of physiological parameters of insect cell cultures during the growth and infection process. Biotechnol Prog 16(5):803–808. https://doi.org/10.1021/bp000092w

    Article  CAS  PubMed  Google Scholar 

  37. Zeiser A, Bdard C, Voyer R et al (1999) On-line monitoring of the progress of infection in Sf-9 insect cell cultures using relative permittivity measurements. Biotechnol Bioeng 63(1):122–126. https://doi.org/10.1002/(SICI)1097-0290(19990405)63:1<122:AID-BIT13>3.0.CO;2-I

    Article  CAS  PubMed  Google Scholar 

  38. Ansorge S, Esteban G, Schmid G (2010) Multifrequency permittivity measurements enable on-line monitoring of changes in intracellular conductivity due to nutrient limitations during batch cultivations of CHO cells. Biotechnol Prog 26(1):272–283. https://doi.org/10.1002/btpr.347

    Article  CAS  PubMed  Google Scholar 

  39. Bluming A, Ziegler J (1971) Regression of Burkitt’s lymphoma in association with measles infection. The Lancet 298(7715):105–106. https://doi.org/10.1016/S0140-6736(71)92086-1

    Article  Google Scholar 

  40. Gopisankar MG, Surendiran A (2018) Oncolytic virotherapy – a novel strategy for cancer therapy. Egyptian J Med Hum Genet 19(3):165–169. https://doi.org/10.1016/j.ejmhg.2017.10.006

    Article  Google Scholar 

  41. Ungerechts G, Bossow S, Leuchs B et al (2016) Moving oncolytic viruses into the clinic: clinical-grade production, purification, and characterization of diverse oncolytic viruses. Mol Ther Methods Clin Dev 3:16018. https://doi.org/10.1038/mtm.2016.18

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Grein TA, Loewe D, Dieken H et al (2018) High titer oncolytic measles virus production process by integration of dielectric spectroscopy as online monitoring system. Biotechnol Bioeng 115(5):1186–1194. https://doi.org/10.1002/bit.26538

    Article  CAS  PubMed  Google Scholar 

  43. Devaux P, von Messling V, Songsungthong W et al (2007) Tyrosine 110 in the measles virus phosphoprotein is required to block STAT1 phosphorylation. Virology 360(1):72–83. https://doi.org/10.1016/j.virol.2006.09.049

    Article  CAS  PubMed  Google Scholar 

  44. Kärber G (1931) Beitrag zur kollektiven Behandlung pharmakologischer Reihenversuche. Archiv f experiment Pathol u Pharmakol 162(4):480–483. https://doi.org/10.1007/BF01863914

    Article  Google Scholar 

  45. Levine DW, Wang DIC, Thilly WG (1979) Optimization of growth surface parameters in microcarrier cell culture. Biotechnol Bioeng 21(5):821–845. https://doi.org/10.1002/bit.260210507

    Article  Google Scholar 

  46. Blüml G (2007) Microcarrier cell culture technology. In: Pörtner R (ed) Animal cell biotechnology, Methods in biotechnology. Humana Press, Totowa, NJ

    Google Scholar 

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Acknowledgments

We thank the Federal Ministry of Education and Research (BMBF) for financial support (Grant No. 13FH001IX5), the Hessen State Ministry of Higher Education, Research and the Arts for financial support within the Hessen initiative for scientific and economic excellence (LOEWE Center for Insect Biotechnology and Bioresources), and Richard M. Twyman for professional editing of the manuscript.

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Authors and Affiliations

  1. Institute of Bioprocess Engineering and Pharmaceutical Technology (IBPT), Technische Hochschule Mittelhessen (THM)—University of Applied Sciences, Giessen, Germany

    Lukas Käßer, Jan Zitzmann, Tanja Grein, Tobias Weidner, Denise Salzig & Peter Czermak

  2. Faculty of Biology and Chemistry, Justus-Liebig-University Giessen, Giessen, Germany

    Peter Czermak

  3. Division Bioresources, Fraunhofer Institute for Molecular Biology and Applied Ecology (IME), Giessen, Germany

    Peter Czermak

Authors
  1. Lukas Käßer
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  2. Jan Zitzmann
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  3. Tanja Grein
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  4. Tobias Weidner
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  5. Denise Salzig
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  6. Peter Czermak
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Corresponding author

Correspondence to Peter Czermak .

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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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Käßer, L., Zitzmann, J., Grein, T., Weidner, T., Salzig, D., Czermak, P. (2020). Turbidimetry and Dielectric Spectroscopy as Process Analytical Technologies for Mammalian and Insect Cell Cultures. 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_20

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  • DOI: https://doi.org/10.1007/978-1-0716-0191-4_20

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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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