Skip to main content
PMC home page
Cytotechnology logoLink to Cytotechnology
. 2006 Jun 23;50(1-3):35–48. doi: 10.1007/s10616-005-3974-x

On-line Measurements and Control of Viable Cell Density in Cell Culture Manufacturing Processes using Radio-frequency Impedance

John P Carvell 1,✉, Jason E Dowd 2
PMCID: PMC3475999  PMID: 19003069

Abstract

In this work, radio-frequency (RF) impedance is reviewed as a method for monitoring and controlling cell culture manufacturing processes. It is clear from the many publications cited that RF Impedance is regarded as an accurate and reliable method for measuring the live cell bio-volume both on-line and off-line and the technology is also sutable for animal cells in suspension, attached to micro-carriers or immobilized in fixed beds. In cGMP production, RF Impedance is being used in three main areas. Firstly, it is being used as a control instrument for maintaining consistent perfusion culture allowing the bioreactor to operate under optimum conditions for maximum production of recombinant proteins. In the second application it has not replaced traditional off-line live cell counting techniques but it is being used as an additional monitoring tool to check product conformance. Finally, RF Impedance is being used to monitor the concentration of live cells immobilized on micro-carriers or packed beds in cGMP processes where traditional off-line live cell counting methods are inaccurate or impossible to perform.

Key words: Radio-frequency impedance, Capacitance, Cell culture, CHO, cGMP, On-line biomass monitoring, Perfusion culture

References

  1. Asami K., Yonezawa T., Wakamatsu H., Koyanagi N. Dielectric spectroscopy of biological cells. Biochem. Bioener. 1996;40:141–145. doi: 10.1016/0302-4598(96)05067-2. [DOI] [Google Scholar]
  2. Barer M.R., Kaprelyants A.S., Weichart D.H., Harwood C.R., Kell D.B. Microbial stress and culturability: conceptual and operational domains. Microbiology UK. 1998;144:2009–2010. doi: 10.1099/00221287-144-8-2009. [DOI] [PubMed] [Google Scholar]
  3. Belving H., Ericksson L.E.G., Davey C.L., Kell D.B. Dielectric properties of human blood and erythrocytes at radio frequencies (0.2–10 MHz); dependence on cell volume fraction and medium composition. Eur. Biophys. J. 1994;23:207–215. doi: 10.1007/BF01007612. [DOI] [PubMed] [Google Scholar]
  4. Carvell J.P. 2003. Monitoring live cell concentration in real time. Bioprocess Int.: 2–7.
  5. Cerkel I., Garcia A., Degouys V., Dubois D., Fabry L., Miller A.O.A. Dielectric-spectroscopy of mammalian cells: evaluation of the biomass of Hela-Cell and CHO-cells in suspension by low frequency dielectric spectroscopy. Cytotechnology. 1993;13:185–193. doi: 10.1007/BF00749814. [DOI] [PubMed] [Google Scholar]
  6. Clegg J.S. Properties and metabolism of the aqueous cytoplasm and its boundaries. Am. J. Physiol. 1984;246:R133–R151. doi: 10.1152/ajpregu.1984.246.2.R133. [DOI] [PubMed] [Google Scholar]
  7. Davey C.L. and Kell D.B. 1995. The low-frequency dielectric properties of biological cells. In: Bioelectrochemistry: Principles and PracticeVol. 2. Bioelectrochem’of Cells and Tissues. pp. 159–207.
  8. Davey C.L., Davey H.M., Kell D.B., Todd R.W. Introduction to the dielectric estimation of cellular biomass in real timewith special emphasis on measurements at high volume fractions. Anal. Chim. Acta. 1993a;279:155–161. doi: 10.1016/0003-2670(93)85078-X. [DOI] [Google Scholar]
  9. Davey C.L., Markx G.H., Kell D.B. On the dielectric method of measuring cellular viability. Pure Appl. Chem. 1993b;65:1921–1926. doi: 10.1351/pac199365091921. [DOI] [Google Scholar]
  10. Davey C.L., Guan Y., Kemp R.B. Real time monitoring of the biomass content of animal cell cultures using dielectric spectroscopy. Animal Cell Technol.: Basic Appl. Aspects. 1997a;8:61–65. [Google Scholar]
  11. Davey C.L., Guan Y., Kemp R.B., Kell D.B. Real-time monitoring of the biomass content of animal cell cultures using dielectric spectroscopy. In: Funatsu K., Shirai Y., Matsushita T., editors. Animal Cell Technology: Basic and Applied Aspects, Vol. 8. Dordrecht: Kluwer; 1997b. pp. 61–65. [Google Scholar]
  12. Degouys V., Cerkel I., Garcia A., Harfield J., Dubois D., Fabry L., Miller A.O.A. Dielectric spectroscopy of mammalian cells: 2, simultaneous in situ evaluation by aperture impedance pulse spectroscopy and low-frequency dielectric spectroscopy of the biomass of HTC cells on Cytodex 3. Cytotechnology. 1993;13:195–202. doi: 10.1007/BF00749815. [DOI] [PubMed] [Google Scholar]
  13. Dowd J.E., Jubb A., Kwok E.K., Piret J.M. Optimization and control of perfusion cultures using a viable cell probe and cell specific perfusion rates. Cytotechnology. 2003;42:35–45. doi: 10.1023/A:1026192228471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dowd J.E., Carvell J.P. Improved control of cGMP fermentations and cell culture. Genet. Engineer. News. 2005;25(11):64–68. [Google Scholar]
  15. Ducommun P., Kadori A., Von Stockar U., Marison I. On-line determination of animal cell concentration in two industrial high density culture processes by dielectric spectroscopy. Biotechnol. Bioeng. 2002a;77:316–323. doi: 10.1002/bit.1197. [DOI] [PubMed] [Google Scholar]
  16. Ducommun P., Ruffieux P.A., Kadouri A., Stockar U., Marison Monitoring temperature effects on cell metabolism in a packed bed process. Biotechnol. Bioeng. 2002b;77:838–842. doi: 10.1002/bit.10185. [DOI] [PubMed] [Google Scholar]
  17. Elias C.B., Zeiser A., Bedard C., Kamen A.A. Enhanced growth of Sf-9 cells to a maximum density of 5.2×107 cells per ml and production of B-Galactosidase at high cell density by fed batch culture. Biotechnol. Bioeng. 2000;68:381–388. doi: 10.1002/(SICI)1097-0290(20000520)68:4<381::AID-BIT3>3.0.CO;2-D. [DOI] [PubMed] [Google Scholar]
  18. Elias C.B., Zeiser A., Kamen A.A. Advances in high cell density culture technology using the Sf-9 insect cell baculovirus expression system — the fed batch approach. Bioprocess J. 2003;2(1):22–29. [Google Scholar]
  19. Ferreira A.P., Vieira L.M., Cordoso J.P., Menzes J.C. Evaluation of a new capacitance probe for biomass monitoring in industrial pilot-scale fermentations. J. Biotechnol. 2005;116:403–409. doi: 10.1016/j.jbiotec.2004.12.006. [DOI] [PubMed] [Google Scholar]
  20. Foster K.R., Schwan H.P. Dielectric properties of tissues. In: Polk C., Postow E., editors. CRC Handbook of Biological Effects of Electromagnetuic Fields. Boca Raton, FL: CRC Press; 1986. [Google Scholar]
  21. Foster K.R., Schwan H.P. Dielectric properties of tissues and biological materials: a critical review. Critical Reviews in Biomedical Engineering. 1989;17:25–104. [PubMed] [Google Scholar]
  22. Guan Y., Kemp R. The viable cell monitor: a dielectric spectroscope for growth and metabolic studies of animal cells on macroporous beads. In: Merten O.-W., Perrin P., Griffiths B., editors. New Developments and New Applications in Animal Cell Technology. Dordrecht/NL: Springer Netherlands; 1997. pp. 321–328. [Google Scholar]
  23. Guan Y., Evans P.M., Kemp R.B. An on-line monitor and potential control variable of specific metabolic rate in animal cell culture that combines microcalorimetry with dielectric spectroscopy. Biotechnol. Bioeng. 1998;58:463–477. doi: 10.1002/(SICI)1097-0290(19980605)58:5<464::AID-BIT2>3.0.CO;2-B. [DOI] [PubMed] [Google Scholar]
  24. Harris C.M., Todd R.W., Bungard S.H., Lovitt R.W., Morris J.G., Kell D.B. The dielectric permietivity of microbial suspensions at radio frequencies: a novel method for the estimation of microbial biomass. Enzyme Microb. Technol. 1987;9:181–186. doi: 10.1016/0141-0229(87)90075-5. [DOI] [Google Scholar]
  25. Kell D.B., Todd R.W. Dielectric estimation of microbial biomass using the Aber Instruments Biomass Monitor. TIBTECH. 1998;16:149–150. [Google Scholar]
  26. Kell D.B., Markx G.H., Davey C.L., Todd R.W. Real time monitoring of cellular biomass: methods and applications. Trends Anal. Chem. 1990;9:190–194. doi: 10.1016/0165-9936(90)87042-K. [DOI] [Google Scholar]
  27. Kell D.B., Kaprelyants A.S., Weichart D.H., Harwood C.L., Baxter M.R. Viability and activity in readily culturable bacteria: a review and discussion of the practical issues. Antonie van Leeuwenhoek. 1998;73:169–187. doi: 10.1023/A:1000664013047. [DOI] [PubMed] [Google Scholar]
  28. Konstantinov K.B., Pambayun R., Matanguihan R., Yoshida T., Perusich C.M., Hu W.S. On-line monitoring of hybridoma cell growth using a laser turbidity sensor. Biotechnol. Bioeng. 1992;40:1337–1342. doi: 10.1002/bit.260401107. [DOI] [PubMed] [Google Scholar]
  29. Konstantinov K., Chuppa S., Saja E., Tsal Y., Yoons, Golini F. Real time biomass concentration monitoring in animal cell cultures. TIBTECH. 1994;12:324–333. doi: 10.1016/0167-7799(94)90049-3. [DOI] [PubMed] [Google Scholar]
  30. Merten O.-W., Palfi G.E., Stäheli J., Steiner J. Invasive infrared sensor for the determination of the cell number in a continuous fermentation of hybridomas. Dev. Biol. Standard. 1987;66:357–360. [PubMed] [Google Scholar]
  31. Noll T., Biselli M. Dielectric spectroscopy in the cultivation of suspended and immobilised hybridoma cells. J. Biotechnol. 1998;63:187–198. doi: 10.1016/S0168-1656(98)00080-7. [DOI] [PubMed] [Google Scholar]
  32. Pethig R. Dielectric and Electronic Properties of Biological Materials. Chichester: Wiley; 1979. [Google Scholar]
  33. Pethig R., Kell D.B. The passive electrical properties of biological systems: their significance in physiology, biophysics and biotechnology. Phys. Med. Biol. 1987;32:933–970. doi: 10.1088/0031-9155/32/8/001. [DOI] [PubMed] [Google Scholar]
  34. Schmid G., Zacher D. Evaluation of a novel capacitance probe for on-line monitoring of viable cell densities in batch and fed-bach animal cell culture processes. In: Godia F., Fussenegger M., editors. Animal Cell Technology Meets Genomics. Dordrecht/NL: Springer; 2004. pp. 621–624. [Google Scholar]
  35. Siano S.A. Biomass measurement by inductive permeattivity. Biotechnol. Bioeng. 1997;55:289–304. doi: 10.1002/(SICI)1097-0290(19970720)55:2<289::AID-BIT7>3.0.CO;2-E. [DOI] [PubMed] [Google Scholar]
  36. Stoicheva N.G., Davey C.L., Markx G.H., Kell D.B. Dielectric spectroscopy: a rapid method for the determination of solvent biocompatibility during biotransformations. Biocatalysis. 1989;2:5–22. doi: 10.3109/10242428908992034. [DOI] [Google Scholar]
  37. Takashima S., Asami K., Takahashi Y. Frequency domain studies of impedance characteristics of biological cells using micropippette technique. 1. Erythrocyte. Biophys. J. 1988;54:995–1000. doi: 10.1016/S0006-3495(88)83037-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Vits H., Hu W.S. Fluctuations in continuous mammalian cell bioreactors with retention. Biotechnol. Progr. 1992;8:397–403. doi: 10.1021/bp00017a004. [DOI] [PubMed] [Google Scholar]
  39. Wu P., Ozturk S., Blackie J.D., Thrift J.C., Figueroa C., Naveh D. Evaluation and applications of optical density probes in mammalian cell bioreactors. Biotechnol. Bioeng. 1995;45:495–502. doi: 10.1002/bit.260450606. [DOI] [PubMed] [Google Scholar]
  40. Zeiser A., Bedard C., Voyer R., Jardin B., Tom R., Karmen A.A., Karmen T. On-line monitoring of the progress of infection in Sf-9 insect cell cultures using relative permittivity measurements. Biotechnol. Bioeng. 1999;63:122–126. doi: 10.1002/(SICI)1097-0290(19990405)63:1<122::AID-BIT13>3.0.CO;2-I. [DOI] [PubMed] [Google Scholar]
  41. Zeiser A., Voyer R., Jardin B., Kamen A. On-line monitoring physiological parameters of insect cell cultures during growth and infection process. Biotechnol. Progr. 2000;16:803–808. doi: 10.1021/bp000092w. [DOI] [PubMed] [Google Scholar]
  42. Zho W., Hu W.S. On-line characterisation of a hybridoma cell culture process. Biotechnol. Bioeng. 1994;44:170–177. doi: 10.1002/bit.260440205. [DOI] [PubMed] [Google Scholar]

Articles from Cytotechnology are provided here courtesy of Springer Science+Business Media B.V.

RESOURCES