Skip to main content
SpringerLink
Log in

Adaptation of the Spodoptera exigua Se301 insect cell line to grow in serum-free suspended culture. Comparison of SeMNPV productivity in serum-free and serum-containing media

  • Biotechnological products and process engineering
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Spodoptera exigua Se301 cells have been successfully adapted to two different commercial serum-free media (SFM; Ex-Cell 420 and Serum-Free Insect Medium-1) by gradually reducing the 10 %-added serum-containing medium content from 100 % to 0 % (v/v) in suspended cultures. Both direct adaptation to a serum-free medium and cell growth in the absence of protective additives against fluid dynamic stress [polyvinyl pyrrolidone and polyvinyl alcohol] and disaggregation [dextran sulfate] proved impossible. Cells grew reproducibly in both SFMs once the serum had been completely removed, although the use of Ex-Cell 420 resulted in higher growth rates and cell densities. Turbulence was sufficiently high to reduce growth rates and final cell densities at the highest Reynolds number investigated, although no clear influence of agitation was observed on virus productivity. Both attached and suspended Se301 cell cultures were successfully infected with the SeMNPV baculovirus. Cells adapted to different conditions (attached or suspended culture, serum-containing or serum-free medium) showed different occlusion bodies productivities at the two multiplicities of infection assayed (0.1 and 0.5).

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  • Akhoukh R, Kretzmer G, Schügerl K (1996) On-line monitoring and control of the cultivation of Spodoptera frugiperda Sf9 insect cells and β-galactosidase production by Autographa californica virus vector. Enzyme Microb Technol 18:220–228

    Article  Google Scholar 

  • Beas-Catena A, Sánchez-Mirón A, García-Camacho F, Molina-Grima E (2011) Adaptation of the Se301 insect cell line to suspension culture. Effect of turbulence on growth and on production of nucleopolyhedrovius (SeMNPV). Cytotechnology 63:543–552

    Article  CAS  Google Scholar 

  • Bonning BC, Hoover K, Duffey S, Hammock BD (1995) Production of polyhedra of the Autographa californica nuclear polyhedrosis virus using the Sf21 and Tn5B1-4 cell lines and comparison with host-derived polyhedra by bioassay. J Invert Pathol 66:224–230

    Article  CAS  Google Scholar 

  • Chakraborty S, Monsour C, Teakle R, Reid S (1999) Yield, biological activity, and field performance of a wild-type Helicoverpa nucleopolyhedrovirus produced in H. zea cell cultures. J Invert Pathol 73:199–205

    Article  CAS  Google Scholar 

  • Chen Q, McIntosh AH, Yu Z, Hong H, Goodman CL, Grasela JJ, Ignoffo CM (1993) The replication of Autographa californica baculovirus (AcMNPV) in 2 lepidopteran cell lines grown in serum-free media. J Invert Pathol 62:216–219

    Article  Google Scholar 

  • Claus JD, Remondetto GE, Guerrero SA, Demonte AM, Murguía M, Marcipar AJ (1993) Anticarsia gemmatalis nuclear polyhedrosis virus replication in serum-free and serum-reduced insect cell cultures. J Biotechnol 31:1–15

    Article  CAS  Google Scholar 

  • Cruz PE, Moreira JL, Larrondo MJT (1997) Insect cell growth evaluation during serum-free adaptation in stirred suspension cultures. Biotechnol Tech 11(2):117–120

    Article  CAS  Google Scholar 

  • Cruz PE, Cunha A, Peixoto CC, Clemente J, Moreira JL, Carrondo MJT (1998) Optimization of the production of virus-like particles in insect cells. Biotechnol Bioeng 60:408–418

    Article  CAS  Google Scholar 

  • Dee KU, Shuler ML (1997) Optimization of an assay for baculovirus titer and design of regimens for the synchronous infection of insect cells. Biotechnol Prog 13:14–24

    Article  CAS  Google Scholar 

  • Dee KU, Shuler ML, Wood HA (1997a) Inducing single-cell suspension of BTI-TN5B1-4 insect cells. I The use of sulfated polyanions to prevent cell aggregation and enhance recombinant protein production. Biotechnol Bioeng 54:191–205

    Article  CAS  Google Scholar 

  • Dee KU, Wood HA, Shuler ML (1997b) Inducing single-cell suspension of BTI-TN5B1-4 insect cells. II The effect of sulfated polyanions on baculovirus infection. Biotechnol Bioeng 54:206–220

    Article  CAS  Google Scholar 

  • García Camacho F, Gallardo Rodríguez JJ, Sánchez Mirón A, Cerón García MC, Belarbi EH, Molina Grima E (2007) Determination of shear stress thresholds in toxic dinoflagellates cultured in shaken flasks. Implications in bioprocess engineering. Process Biochem 42:1506–1515

    Article  Google Scholar 

  • Gelernter WD, Federici BA (1986) Continuous cell line from Spodoptera exigua (Lepidoptera: Noctuidae) that supports replication of nuclear polyhedrosis viruses from Spodoptera exigua and Autographa californica. J Invert Pathol 48:199–207

    Article  Google Scholar 

  • Goldblum S, Bae YK, Hink WF, Chalmers J (1990) Protective effect of methylcellulose and other polymers on insect cells subjected to laminar shear stress. Biotechnol Prog 6:383–390

    Article  CAS  Google Scholar 

  • Goodman CL, El Sayed GN, McIntosh AH, Grasela JJ, Stilles B (2001) Establishment and characterization of insect cell lines from 10 lepidopteran species. In Vitro Cell Dev Biol Anim 37:367–373

    CAS  Google Scholar 

  • Hara K, Funakoshi M, Tsuda K, Kawarabata T (1993) New Spodoptera exigua cell lines susceptible to Spodoptera exigua nuclear polyhedrosis virus. In Vitro Cell Dev Biol Anim 29:904–907

    Article  Google Scholar 

  • Hink WF, Strauss EM, Ramoska WA (1977) Propagation of Autographa californica nuclear polyhedrosis virus in cell culture: methods for infecting cells. J Invert Pathol 30:185–191

    Article  Google Scholar 

  • Ikonomou L, Bastin G, Schneider YJ, Agathos SN (2004) Effect of partial medium replacement on cell growth and protein production for the high-five™ insect cell line. Cytotechnology 44:67–76

    Article  CAS  Google Scholar 

  • Jardin BA, Montes J, Lanthier S, Tran R, Elías C (2007) High cell density fed batch and perfusion processes for stable non-viral expression of secreted alkaline phosphatase (SEAP) using insect cells: comparison to a batch Sf-9-BEV system. Biotechnol Bioeng 97:332–345

    Article  CAS  Google Scholar 

  • King LA, Possee RD (1992) The baculovirus expression system. Chapman & Hall, London

    Book  Google Scholar 

  • Kunas KT, Papoutsakis ET (1989) Increasing serum concentrations decrease cell death and allow growth of hybridoma cells at higher agitation rates. Biotechnol Lett 8:525–530

    Article  Google Scholar 

  • Marteijn RCL, Jurrius O, Dhont J, De Gooijier CD, Tramper J, Martens DE (2003) Optimization of a feed medium for fed-batch culture of insect cells using a genetic algorithm. Biotechnol Bioeng 81:269–278

    Article  CAS  Google Scholar 

  • Michaels JD, Nowak JE, Mallik AK, Koczo K, Wasan DT, Papoutsakis ET (1995) Analysis of cell-to-bubble attachment in sparged bioreactors in the presence of cell-protecting additives. Biotechnol Bioeng 47:407–419

    Article  CAS  Google Scholar 

  • Micheloud GA, Gioria VV, Pérez G, Claus JD (2009) Production of occlusion bodies of Anticarsia gemmatalis multiple nucleopolyhedrovirus in serum-free suspension cultures of the saUFL-AG-286 cell line. Influence of infection conditions and statistical optimization. J Virol Methods 162:258–266

    Article  CAS  Google Scholar 

  • Palomares LA, González M, Ramírez OT (2000) Evidence of Pluronic F-68 direct interaction with insect cells: impact on shear protection, recombinant protein, and baculovirus production. Enzyme Microb Technol 26:324–331

    Article  CAS  Google Scholar 

  • Papoutsakis ET (1991) Media additives for protecting freely suspended animal cells against agitation and aeration damage. Trends Biotechnol 9:316–324

    Article  CAS  Google Scholar 

  • Petricevich VL, Palomares LA, González M, Ramírez OT (2001) Parameters that determine virus adsorption kinetics: toward the design of better infection strategies for the insect cell—baculovirus expression system. Enzyme Microb Technol 29:56–61

    Article  Google Scholar 

  • Radford KM, Reid S, Greenfield PF (1997a) Substrate limitation in the baculovirus expression vector system. Biotechnol Bioeng 56:32–44

    Article  CAS  Google Scholar 

  • Radford KM, Cavegn C, Bertrand M, Bernard AR, Reid S, Greenfield PF (1997b) The indirect effects of multiplicity of infection on baculovirus expressed proteins in insect cells: secreted and non-secreted products. Cytotechnology 24:73–81

    Article  CAS  Google Scholar 

  • Rhiel M, Mitchell-Logean CM, Murhammer DW (1997) Comparison of Trichoplusia ni BTI-Tn-5b1-4 (high fiveTM) and Spodoptera frugiperda Sf-9 insect cell line metabolism in suspension cultures. Biotechnol Bioeng 55:909–920

    Article  CAS  Google Scholar 

  • Rodas VM, Marques FH, Honda MT, Soares DM, Jorge SA, Antoniazzi MM, Medugno C, Castro ME, Ribeiro BM, Souza ML, Tonso A, Pereira CA (2005) Cell culture derived AgMNPV bioinsecticide: biological constraints and bioprocess issues. Cytotechnology 48:27–39

    Article  CAS  Google Scholar 

  • Sugiura T, Amann E (1996) Properties of two insect cell lines useful for the baculovirus expression system in serum-free culture. Biotechnol Bioeng 51:494–499

    Article  CAS  Google Scholar 

  • Taticek RA, Choi C, Phan SE, Palomares LA, Shule ML (2001) Comparison of growth and recombinant protein expression in two different insect cell lines in attached and suspension culture. Biotechnol Prog 17:676–684

    Article  CAS  Google Scholar 

  • Tramper J, Williams JB, Joustra D, Vlak JM (1986) Shear sensitivity of insect cells in suspension. Enzyme Microb Technol 8:33–46

    Article  Google Scholar 

  • Vail PV, Jay DL, Hink WF (1973) Replication and infectivity of the nuclear polyhedrosis virus of the alfalfa looper, Autographa californica, produced in cells grown in vitro. J Invert Pathol 22:231–237

    Article  Google Scholar 

  • Wu J, King G, Daugulis AJ, Faulkner P, Bone DH, Goosen MFA (1990) Adaptation of insect cells to suspension culture. J Ferment Bioeng 70(2):90–93

    Article  CAS  Google Scholar 

  • Zhang Z, Chisti Y, Moo-young M (1995) Effects of the hydrodynamic environment and shear protectants on survival of erythrocytes in suspension. J Biotechnol 43:33–40

    Article  CAS  Google Scholar 

  • Zhang YH, Enden G, Merchuk JC (2005) Insect cells–baculovirus system: factors affecting growth and low MOI infection. Biochem Eng J 27:8–16

    Article  CAS  Google Scholar 

  • Zhang H, Zhang YA, Qin Q, Li X, Miao L, Wang Y, Yang Z, Ding C (2006) New cell lines from larval fat bodies of Spodoptera exigua: characterization and susceptibility to baculoviruses (Lepidoptera: Noctuidae). J Invertebr Pathol 91:9–12

    Article  CAS  Google Scholar 

  • Zhang H, Zhang YA, Qin Q, Li X, Miao L, Wang Y, Qu L, Zhang A, Yang Q (2009) A cell strain cloned from Spodoptera exigua cell line (IOZCAS-Spex-II): highly susceptible to S. exigua nucleopolyhedrovirus infection. In Vitro Cell Dev Biol Anim 45:201–204

    Article  Google Scholar 

Download references

Acknowledgments

This work was funded by the Junta de Andalucía, Instituto Andaluz de Biotecnología (BIOANDALUS 0.8/2/L1.2) and Biocolor, S.L.

Author information

Authors and Affiliations

  1. Department of Chemical Engineering, University of Almeria, 04120, Almeria, Spain

    A. Beas-Catena, A. Sánchez-Mirón, F. García-Camacho, A. Contreras-Gómez & E. Molina-Grima

Authors
  1. A. Beas-Catena
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. A. Sánchez-Mirón
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. F. García-Camacho
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. A. Contreras-Gómez
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. E. Molina-Grima
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to A. Sánchez-Mirón.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Beas-Catena, A., Sánchez-Mirón, A., García-Camacho, F. et al. Adaptation of the Spodoptera exigua Se301 insect cell line to grow in serum-free suspended culture. Comparison of SeMNPV productivity in serum-free and serum-containing media. Appl Microbiol Biotechnol 97, 3373–3381 (2013). https://doi.org/10.1007/s00253-012-4576-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-012-4576-0

Keywords

Search

Navigation