Advertisement

Cytotechnology

, Volume 57, Issue 1, pp 37–44 | Cite as

Insect cells respiratory activity in bioreactor

  • Marilena Martins Pamboukian
  • Soraia Athie Calil Jorge
  • Mariza Gerdulo Santos
  • Adriana Yurie Yokomizo
  • Carlos Augusto Pereira
  • Aldo Tonso
Article

Abstract

Specific respiration rate ( \( Q_{{{\text{O}}_{2} }} \)) is a key parameter to understand cell metabolism and physiological state, providing useful information for process supervision and control. In this work, we cultivated different insect cells in a very controlled environment, being able to measure \( Q_{{{\text{O}}_{2} }} \). Spodoptera frugiperda (Sf9) cells have been used through virus infection as host for foreign protein expression and bioinsecticide production. Transfected Drosophila melanogaster (S2) cells can be used to produce different proteins. The objective of this work is to investigate respiratory activity and oxygen transfer during the growth of different insect cells lines as Spodoptera frugiperda (Sf9), Drosophila melanogaster (S2) wild and transfected for the expression of GPV and EGFP. All experiments were performed in a well-controlled 1-L bioreactor, with SF900II serum free medium. Spodoptera frugiperda (Sf9) cells reached 10.7 × 106 cells/mL and maximum specific respiration rate (\( Q_{{{\text{O}}_{2} \max }} \)) of 7.3 × 10−17 molO2/cell s. Drosophila melanogaster (S2) cells achieved 51.2 × 106 cells/mL and \( Q_{{{\text{O}}_{2} \max }} \) of 3.1 × 10–18 molO2/cell s. S2AcGPV (expressing with rabies virus glycoprotein) reached 24.9 × 106 cells/mL and \( Q_{{{\text{O}}_{2} \max }} \) of 1.7 × 10–17 molO2/cell s, while S2MtEGFP (expressing green fluorescent protein) achieved 15.5 × 106 cells/mL and \( Q_{{{\text{O}}_{2} \max }} \) = 1.9 × 10−17 molO2/cell s. Relating to the Sf9, S2 cells reached higher maximum cell concentrations and lower specific respiration rate, which can be explained by its smaller size. These results presented useful information for scale-up and process control of insect cells.

Keywords

Dissolved oxygen Specific respiration rate Drosophila melanogaster S2 Spodoptera frugiperda Sf9 Transfected cell lines 

Notes

Acknowledgments

This work was supported in part by grants from FAPESP (02/09482-3) and CNPQ. The authors thank Marcos Lemos and Alexandra Santos for their collaboration.

References

  1. Bailey JE, Ollis DF (1986) Biochemical engineering fundamentals, 2nd edn. McGraw-Hill, New York, pp 459–469Google Scholar
  2. Chang KH, Yang JM, Chum HOK, Chung IS (2005) Enhance activity of recombinant of β-secretase from Drosophila melanogaster S2 cells transformed with cDNAs encoding human β1,4-galactosytransferase and Gal β1,4-GlcNac α2,6-sialytransferase. J Biotechnol 116:359–367CrossRefGoogle Scholar
  3. Delm L, Wolf H, Wagner R (1999) High level expression of hepatitis B virus surface antigen in stably transfected in Drosophila Shneider-2 cells. J Virol Methods 79:191–203CrossRefGoogle Scholar
  4. Kamen AA, Berdad C, Tom R, Perret S, Jardin B (1996) On-line monitoring of respiration in recombinant-baculovirus infected and uninfected insect cell bioreactor cultures. Biotechnol Bioeng 50:36–48CrossRefGoogle Scholar
  5. Kioukia N, Nienow AW, Emery AN, Albureai M (1995) Physiological and environmental-factors affecting the growth of insect cells and infection with baculovirus. J Biotechnol 38:243–251CrossRefGoogle Scholar
  6. Li B, Tsing S, Kosaka AH, Nguyen B, Osen EG, Bach C, Chan H, Barnett J (1996) Expression of human dopamine β hydroxilase in Drosophila Schneider 2 cells. Biochem J 313:57–64Google Scholar
  7. Mitsuhashi J (1998) Cell culture. In: Hunter-Fujita FR, Entwistle PF, Evans HF, Crook NE (eds) Insect viruses and pest management. John Wiley, England, pp 485–517Google Scholar
  8. Pereira CA, Pouliquen Y, Rodas V, Massotte D, Mortensen C, Sogayar MC, de Murcia J (2001) Optimized insect cell culture for the production of recombinant heterologous proteins and baculovirus particles. BioTechniques 31:1262–1268Google Scholar
  9. Perret BG, Wagner R, Lecat S, Brillet K, Rabut G, Bucher B, Pattus F (2003) Expression EGFP-amino-tagged human mu opioid receptor in Drosophila Schneider 2 cells: a potential expression system for large-scale production of G-protein coupled receptors. Protein Expr Purif 31:123–132CrossRefGoogle Scholar
  10. Rodas VM, Marques FH, Honda MT, Soares DM, Jorge SAC, Antoniazzi MM, Medugno C, Castro MEB, Ribeiro BM, Souza ML, Tonso A, Pereira CA (2005) Cell culture derived AgMNPV bioinsecticide: biological constants and bioprocess issues. Cytotechnology 48:27–39CrossRefGoogle Scholar
  11. Santos MG, Jorge SAC, Brillet K, Pereira CA (2007) Improving heterologus protein expression in transfected Drosophila S2 cells as assessed by EGFP expression. Cytotechnology 54:15–24CrossRefGoogle Scholar
  12. Weber W, Weber E, Geisse S, Memmert K (2002) Optimization of protein expression and establishment of the wave bioreactor for baculovirus/insect cell culture. Cytotechnology 38:77–85CrossRefGoogle Scholar
  13. Weiss SA, Vaughn JL (1986) Cell culture methods for largescale propagation of baculoviruses. In: Granados RR, Federici BA (eds) The biology of baculoviruses, vol II. CRC Press LLC, Boca Raton, pp 63–87Google Scholar
  14. Yokomizo AY, Jorge SAC, Astray RM, Fernandes I, Ribeiro OG, Horton DSP, Tonso A, Tordo N, Pereira CA (2007) Rabies virus glycoprotein expression in Drosophila S2 cells I. Functional recombinant protein in stable co-transfected cell line. Biotechnol J 2:102–109CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Marilena Martins Pamboukian
    • 1
  • Soraia Athie Calil Jorge
    • 1
  • Mariza Gerdulo Santos
    • 1
  • Adriana Yurie Yokomizo
    • 1
  • Carlos Augusto Pereira
    • 1
  • Aldo Tonso
    • 1
  1. 1.Escola Politécnica da Usp - Departamento de Engenharia Química São PauloBrazil

Personalised recommendations