Skip to main content

Transient Gene Expression in Suspension HEK293-EBNA1 Cells

  • Protocol
  • First Online:
Recombinant Protein Expression in Mammalian Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1850))

Abstract

Transient gene expression in human embryo kidney 293 (HEK293) cells is an established approach for the rapid production of large amounts of recombinant proteins (r-proteins). Milligram to gram quantities of r-proteins can be typically obtained within less than 10 days following transfection. In this chapter, we describe a simple and robust transfection process of suspension-growing human embryo kidney 293 cells using two commercially available serum-free media and polyethylenimine as the transfection reagent. This chapter provides examples for the production and purification of a his-tagged recombinant protein and two monoclonal antibodies.

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

Access this chapter

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 99.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 129.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 179.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Lalonde ME, Durocher Y (2017) Therapeutic glycoprotein production in mammalian cells. J Biotechnol 251:128–140

    Article  CAS  Google Scholar 

  2. Durocher Y, Butler M (2009) Expression systems for therapeutic glycoprotein production. Curr Opin Biotechnol 20:700–707

    Article  CAS  Google Scholar 

  3. Pham PL, Kamen A, Durocher Y (2006) Large-scale transfection of mammalian cells for the fast production of recombinant protein. Mol Biotechnol 34:225–237

    Article  CAS  Google Scholar 

  4. Geisse S (2009) Reflections on more than 10 years of TGE approaches. Protein Expr Purif 64:99–107

    Article  CAS  Google Scholar 

  5. Baldi L, Hacker DL, Meerschman C, Wurm FM (2012) Large-scale transfection of mammalian cells. Methods Mol Biol 801:13–26

    Article  CAS  Google Scholar 

  6. Chahal P, Durocher Y, Kamen A (2011) In: Butler M et al (eds) Cell Transfection, in Comprehensive Biotechnology. Elsevier Science & Technology, Amsterdam

    Google Scholar 

  7. Cho MS, Yee H, Brown C, Mei B, Mirenda C, Chan S (2003) Versatile expression system for rapid and stable production of recombinant proteins. Biotechnol Prog 19:229–232

    Article  CAS  Google Scholar 

  8. Ogay ID, Lihoradova OA, Azimova SS, Abdukarimov AA, Slack JM, Lynn DE (2006) Transfection of insect cell lines using polyethylenimine. Cytotechnology 51:89–98

    Article  CAS  Google Scholar 

  9. Wölfel J, Essers R, Bialek C, Hertel S, Scholz-Neumann N, Schiedner G (2011) CAP-T cell expression system: a novel rapid and versatile human cell expression system for fast and high yield transient protein expression. BMC Proc 5(Suppl 8):P133

    Article  Google Scholar 

  10. Mori K, Hamada H, Ogawa T, Ohmuro-Matsuyama Y, Katsuda T, Yamaji H (2017) Efficient production of antibody fab fragment by transient gene expression in insect cells. J Biosci Bioeng 124:221–226

    Article  CAS  Google Scholar 

  11. Shen X, Pitol AK, Bachmann V, Hacker DL, Baldi L, Wurm FM (2015) A simple plasmid-based transient gene expression method using high five cells. J Biotechnol 216:67–75

    Article  CAS  Google Scholar 

  12. Suárez-Patiño SF, Mancini RA, Pereira CA, Suazo CA, Mendonça RZ, Jorge SA (2014) Transient expression of rabies virus glycoprotein (RVGP) in Drosophila melanogaster Schneider 2 (S2) cells. J Biotechnol 192(Pt A):255–262

    Article  Google Scholar 

  13. Shen X, Hacker DL, Baldi L, Wurm FM (2014) Virus-free transient protein production in Sf9 cells. J Biotechnol 171:61–70

    Article  CAS  Google Scholar 

  14. Fischer S, Charara N, Gerber A, Wölfel J, Schiedner G, Voedisch B, Geisse S (2012) Transient recombinant protein expression in a human amniocyte cell line: the CAP-T(R) cell system. Biotechnol Bioeng 109:2250–2261

    Article  CAS  Google Scholar 

  15. Gutierrez-Granados S, Cervera L, Segura Mde L, Wolfel J, Godia F (2016) Optimized production of HIV-1 virus-like particles by transient transfection in CAP-T cells. Appl Microbiol Biotechnol 100:3935–3947

    Article  CAS  Google Scholar 

  16. Delafosse L, Xu P, Durocher Y (2016) Comparative study of polyethylenimines for transient gene expression in mammalian HEK293 and CHO cells. J Biotechnol 227:103–111

    Article  CAS  Google Scholar 

  17. Rajendra Y, Hougland MD, Alam R, Morehead TA, Barnard GC (2015) A high cell density transient transfection system for therapeutic protein expression based on a CHO GS-knockout cell line: process development and product quality assessment. Biotechnol Bioeng 112:977–986

    Article  CAS  Google Scholar 

  18. Daramola O, Stevenson J, Dean G, Hatton D, Pettman G, Holmes W, Field R (2014) A high-yielding CHO transient system: coexpression of genes encoding EBNA-1 and GS enhances transient protein expression. Biotechnol Prog 30:132–141

    Article  CAS  Google Scholar 

  19. Atkinson A, Jack GW (1973) Precipitation of nucleic acids with polyethyleneimine and the chromatography of nucleic acids and proteins on immobilised polyethyleneimine. Biochim Biophys Acta 308:41–52

    Article  CAS  Google Scholar 

  20. Boussif O, Lezoualc'h F, Zanta MA, Mergny MD, Scherman D, Demeneix B, Behr JP (1995) A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: polyethylenimine. Proc Natl Acad Sci U S A 92:7297–7301

    Article  CAS  Google Scholar 

  21. Durocher Y, Perret S, Kamen A (2002) High-level and high-throughput recombinant protein production by transient transfection of suspension-growing human 293-EBNA1 cells. Nucleic Acids Res 30:E9

    Article  Google Scholar 

  22. Davies A, Greene A, Lullau E, Abbott WM (2005) Optimisation and evaluation of a high-throughput mammalian protein expression system. Protein Expr Purif 42:111–121

    Article  CAS  Google Scholar 

  23. Abbott WM, Middleton B, Kartberg F, Claesson J, Roth R, Fisher D (2015) Optimisation of a simple method to transiently transfect a CHO cell line in high-throughput and at large scale. Protein Expr Purif 116:113–119

    Article  CAS  Google Scholar 

  24. Soulez M, Sirois I, Brassard N, Raymond MA, Nicodème F, Noiseux N, Durocher Y, Pshezhetsky AV, Hébert MJ (2010) Epidermal growth factor and perlecan fragments produced by apoptotic endothelial cells co-ordinately activate ERK1/2-dependent antiapoptotic pathways in mesenchymal stem cells. Stem Cells 28:810–820

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Yves Durocher .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2018 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

L’Abbé, D., Bisson, L., Gervais, C., Grazzini, E., Durocher, Y. (2018). Transient Gene Expression in Suspension HEK293-EBNA1 Cells. In: Hacker, D. (eds) Recombinant Protein Expression in Mammalian Cells. Methods in Molecular Biology, vol 1850. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8730-6_1

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-8730-6_1

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8729-0

  • Online ISBN: 978-1-4939-8730-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics