Skip to main content
SpringerLink
Log in

Cell Xpress™ Applications in Development and Characterization of Biopharmaceutical Recombinant Protein Producing Cell Lines

  • Chapter
  • First Online:
Cell Line Development

Part of the book series: Cell Engineering ((CEEN,volume 6))

Abstract

The biopharmaceutical industry is focused on the development of quality processes for producing therapeutic proteins and monoclonal antibodies in mammalian cells. The first step in the process development workflow is cell line development. This chapter focuses on applications developed on the LEAPTM (Laser-Enabled Analysis and Processing) instrument to expedite the cell line development process. Applications reviewed include expression optimization, high throughput single cell clone isolation and cell population characterization.

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

Access this chapter

Log in via an institution
Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
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

References

  • Barnes LM, Bentley CM, Dickson AJ (2001) Characterization of the stability of recombinant protein production in the GS-NS0 expression system. Biotechnol Bioeng 73:261–270

    Article  PubMed  CAS  Google Scholar 

  • Barnes L.M, Bentley C.M, Dickson A.J. (2003) Stability of recombinant protein production in the GS-NS0 expression system is unaffected by cryopreservation. Biotechnol Prog 19:233–237

    Article  PubMed  CAS  Google Scholar 

  • Barnes L.M, Bentley C.M, Dickson A.J. (2004) Molecular definition of predictive indicators of stable protein expression in recombinant NS0 myeloma cells. Biotechnol Bioeng 85:115–121

    Article  PubMed  CAS  Google Scholar 

  • Barnes L.M, Moy N, Dickson A.J. (2006) Phenotypic variation during cloning procedures: analysis of the growth behavior of clonal cell lines. Biotechnol Bioeng 94:530–537

    Article  PubMed  CAS  Google Scholar 

  • Barnes L.M, Bentley C.M, Moy N, Dickson A.J (2007) Molecular analysis of successful cell line selection in transfected GS-NS0 myeloma cells. Biotechnol Bioeng 96:337–348

    Article  PubMed  CAS  Google Scholar 

  • Bi J.X, Shuttleworth J, Al-Rubeai M (2004) Uncoupling of cell growth and proliferation results in enhancement of productivity in p21CIP1-arrested CHO cells. Biotechnol Bioeng 85:741–749

    Article  PubMed  CAS  Google Scholar 

  • Bohm E, Voglauer R, Steinfellner W, Kunert R, Borth N, Katinger H (2004) Screening for improved cell performance: selection of subclones with altered production kinetics or improved stability by cell sorting. Biotechnol Bioeng 88:699–706

    Article  PubMed  CAS  Google Scholar 

  • Borth N, Zeyda M, Kunert R, Katinger H (2000) Efficient selection of high-producing subclones during gene amplification of recombinant Chinese hamster ovary cells by flow cytometry and cell sorting. Biotechnol Bioeng 71:266–273

    Article  PubMed  CAS  Google Scholar 

  • Browne S.M, Al-Rubeai M (2007) Selection methods for high-producing mammalian cell lines. Trends Biotechnol 25:425–432

    Article  PubMed  CAS  Google Scholar 

  • Carroll S, Al-Rubeai M (2004) The selection of high-producing cell lines using flow cytometry and cell sorting. Expert Opin Biol Ther 4:1821–1829

    Article  PubMed  CAS  Google Scholar 

  • Carton J.M, Sauerwald T, Hawley-Nelson P, Morse B, Peffer N, Beck H, Lu J, Cotty A, Amegadzie B, Sweet R (2007) Codon engineering for improved antibody expression in mammalian cells. Protein Expr Purif 55:279–286

    Article  PubMed  CAS  Google Scholar 

  • Charlet M, Kromenaker SJ, Srienc F (1995) Surface IgG content of murine hybridomas: direct evidence for variation of antibody secretion rates during the cell cycle. Biotechnol Bioeng 47:535–540

    Article  PubMed  CAS  Google Scholar 

  • Chen R, Greene E.L, Collinsworth G, Grewal J.S, Houghton O, Zeng H, Garnovskaya M, Paul R.V, Raymond J.R. (1999) Enrichment of transiently transfected mesangial cells by cell sorting after cotransfection with GFP. Am J Physiol 276:F777–F785

    PubMed  CAS  Google Scholar 

  • Chenuet S, Martinet D, Besuchet-Schmutz N, Wicht M, Jaccard N, Bon AC, Derouazi M, Hacker D.L, Beckmann J.S, Wurm F.M. (2008) Calcium phosphate transfection generates mammalian recombinant cell lines with higher specific productivity than polyfection. Biotechnol Bioeng 101:937–945

    PubMed  Google Scholar 

  • Coller H.A, Coller BS (1983) Statistical analysis of repetitive subcloning by the limiting dilution technique with a view toward ensuring hybridoma monoclonality. Hybridoma 2:91–96

    Article  PubMed  CAS  Google Scholar 

  • Coller H.A, Coller B.S (1986) Poisson statistical analysis of repetitive subcloning by the limiting dilution technique as a way of assessing hybridoma monoclonality. Methods Enzymol 121:412–417

    Article  PubMed  CAS  Google Scholar 

  • Corisdeo S, Cassel M.J, Kinney C.S, Ganguly S, Kraichely K.M. Use of flow cytometry to screen and predict stability of candidate manufacturing cell lines. 2008; Conference Presentation. Bioprocess International Annual Meeting. Anaheim, CA.

    Google Scholar 

  • Davis L. Optimization of electroporation and clone selection of EB14 chicken embryonic stem cells to express recombinant monoclonal antibodies. 2007; Conference Poster. IBC Cell Line Development and Engineering. San Diego, CA.

    Google Scholar 

  • Dean P.N, Hoffman R.A (2007) Overview of flow cytometry instrumentation. Curr Protoc Cytom Chapter 1: Volume 1 Unit1 1.

    Google Scholar 

  • DeMaria C.T, Cairns V, Schwarz C, Zhang J, Guerin M, Zuena E, Estes S, Karey K.P.(2007) Accelerated clone selection for recombinant CHO CELLS using a FACS-based high-throughput screen. Biotechnol Prog 23:465–472

    Article  PubMed  CAS  Google Scholar 

  • Derouazi M, Flaction R, Girard P, de Jesus M, Jordan M, Wurm F.M. (2006) Generation of recombinant Chinese hamster ovary cell lines by microinjection. Biotechnol Lett 28:373–382

    Article  PubMed  CAS  Google Scholar 

  • Dinnis D.M, James D.C. (2005) Engineering mammalian cell factories for improved recombinant monoclonal antibody production: lessons from nature? Biotechnol Bioeng 91:180–189

    Article  PubMed  CAS  Google Scholar 

  • Fuller S.A, Takahashi M, Hurrell J.G. (2001) Cloning of hybridoma cell lines by limiting dilution. Curr Protoc Mol Biol Chapter 11: http://mrw.interscience.wiley.com/emrw/9780471142720/cp/cpmb/toc Unit11 8.

  • Galfre G, Milstein C (1981) Preparation of monoclonal antibodies: strategies and procedures. Methods Enzymol 73:3–46

    Article  PubMed  CAS  Google Scholar 

  • Hanania EG, Fieck A, Stevens J, Bodzin L.J, Palsson B.O, Koller M.R. (2005) Automated in situ measurement of cell-specific antibody secretion and laser-mediated purification for rapid cloning of highly-secreting producers. Biotechnol Bioeng 91:872–876

    Article  PubMed  CAS  Google Scholar 

  • Hinterkorner G, Brugger G, Muller D, Hesse F, Kunert R, Katinger H, Borth N (2007) Improvement of the energy metabolism of recombinant CHO cells by cell sorting for reduced mitochondrial membrane potential. J Biotechnol 129:651–657

    Article  PubMed  CAS  Google Scholar 

  • Holliger P, Hudson P.J. (2005) Engineered antibody fragments and the rise of single domains. Nat Biotechnol 23:1126–1136

    Article  PubMed  CAS  Google Scholar 

  • Jiang Z, Sharfstein S.T. (2008) Sodium butyrate stimulates monoclonal antibody over-expression in CHO cells by improving gene accessibility. Biotechnol Bioeng 100:189–194

    Article  PubMed  CAS  Google Scholar 

  • Jiang Z, Huang Y, Sharfstein S.T. (2006) Regulation of recombinant monoclonal antibody production in chinese hamster ovary cells: a comparative study of gene copy number, mRNA level, and protein expression. Biotechnol Prog 22:313–318

    Article  PubMed  CAS  Google Scholar 

  • Jun S.C, Kim M.S, Hong H.J, Lee G.M. (2006) Limitations to the development of humanized antibody producing Chinese hamster ovary cells using glutamine synthetase-mediated gene amplification. Biotechnol Prog 22:770–780

    Article  PubMed  CAS  Google Scholar 

  • Kalwy S, Rance J, Young R (2006) Toward more efficient protein expression: keep the message simple. Mol Biotechnol 34:151–156

    Article  PubMed  CAS  Google Scholar 

  • Kim C.H, Oh Y, Lee T.H. (1997) Codon optimization for high-level expression of human erythropoietin (EPO) in mammalian cells. Gene 199:293–301

    Article  PubMed  CAS  Google Scholar 

  • Kim N.S, Byun T.H, Lee G.M. (2001) Key determinants in the occurrence of clonal variation in humanized antibody expression of cho cells during dihydrofolate reductase mediated gene amplification. Biotechnol Prog 17:69–75

    Article  PubMed  CAS  Google Scholar 

  • Koller M.R, Hanania E.G, Stevens J, Eisfeld T.M, Sasaki G.C, Fieck A, Palsson B.O. (2004) High-throughput laser-mediated in situ cell purification with high purity and yield. Cytometry A 61:153–161

    Article  PubMed  Google Scholar 

  • Kromenaker S.J, Srienc F (1994) Stability of producer hybridoma cell lines after cell sorting: a case study. Biotechnol Prog 10:299–307

    Article  PubMed  CAS  Google Scholar 

  • Lin N, Cresswell J.R, Richardson G.A, Gerber M.A, Kayser K.J (2008) Methods and applications of laser-enabled analysis and processing (LEAP). Curr Protoc Cytom Chapter 2: Volume 1 Unit2 14.

    Google Scholar 

  • Liu C, Dalby B, Chen W, Kilzer J.M, Chiou H.C. (2008) Transient transfection factors for high-level recombinant protein production in suspension cultured mammalian cells. Mol Biotechnol 39:141–153

    Article  PubMed  CAS  Google Scholar 

  • Mazur X, Fussenegger M, Renner W.A, Bailey J.E. (1998) Higher productivity of growth-arrested Chinese hamster ovary cells expressing the cyclin-dependent kinase inhibitor p27. Biotechnol Prog 14:705–713

    Article  PubMed  CAS  Google Scholar 

  • Meents H, Enenkel B, Werner R.G, Fussenegger M (2002) p27Kip1-mediated controlled proliferation technology increases constitutive sICAM production in CHO-DUKX adapted for growth in suspension and serum-free media. Biotechnol Bioeng 79:619–627

    Article  PubMed  CAS  Google Scholar 

  • Park S.H, Ryu D.D (1994) Cell cycle kinetics and monoclonal antibody productivity of hybridoma cells during perfusion culture. Biotechnol Bioeng 44:361–367

    Article  PubMed  CAS  Google Scholar 

  • Porter A, Barnes L, Dickson A, Rancher A. Behaviour of GS-CHO cell lines in a selection strategy. 2007; Conference Presentation. ESACT 2007. Dresden, Germany.

    Google Scholar 

  • Reichert J.M, Rosensweig C.J, Faden L.B, Dewitz M.C (2005) Monoclonal antibody successes in the clinic. Nat Biotechnol 23:1073–1078

    Article  PubMed  CAS  Google Scholar 

  • Richardson G.A. An evaluation of the intrinsic IgG production capabilities of different Chinese hamster ovary parental cell lines. 2007; Conference Presentation. ESACT 2007. Dresden, Germany.

    Google Scholar 

  • Schreiner C.L, Bauer J.S, Danilov Y.N, Hussein S, Sczekan M.M, Juliano R.L. (1989) Isolation and characterization of Chinese hamster ovary cell variants deficient in the expression of fibronectin receptor. J Cell Biol 109:3157–3167

    Article  PubMed  CAS  Google Scholar 

  • Seifert D.B, Phillips J.A. (1999) The production of monoclonal antibody in growth-arrested hybridomas cultivated in suspension and immobilized modes. Biotechnol Prog 15:655–666

    Article  PubMed  CAS  Google Scholar 

  • Shapiro H.M. (2003) Practical flow cytometry. Chapter 1. Overture. Wiley, Hoboken, NJ, pp 170–171

    Google Scholar 

  • Stein L.D, Ledgley C.J, Sigal N.H. (1983) Patterns of isotype commitment in human B cells: limiting dilution analysis of Epstein Barr virus-infected cells. J Immunol 130:1640–1645

    PubMed  CAS  Google Scholar 

  • Swiderek H, Al-Rubeai M (2007) Functional genome-wide analysis of antibody producing NS0 cell line cultivated at different temperatures. Biotechnol Bioeng 98:616–630

    Article  PubMed  CAS  Google Scholar 

  • Tobey R.A, Oishi N, Crissman H.A.(1990) Cell cycle synchronization: reversible induction of G2 synchrony in cultured rodent and human diploid fibroblasts. Proc Natl Acad Sci USA 87:5104–5108

    Article  PubMed  CAS  Google Scholar 

  • Twyman R.M. (2005) Gene transfer to animal cells. BIO Scientific. Oxford, UK. pp 1–18.

    Google Scholar 

  • Underwood P.A, Bean P.A. (1988) Hazards of the limiting-dilution method of cloning hybridomas. J Immunol Methods 107:119–128

    Article  PubMed  CAS  Google Scholar 

  • Wurm F.M. (2004) Production of recombinant protein therapeutics in cultivated mammalian cells. Nat Biotechnol 22:1393–1398

    Article  PubMed  CAS  Google Scholar 

  • Wurm F.M, Bernard A (1999) Large-scale transient expression in mammalian cells for recombinant protein production. Curr Opin Biotechnol 10:156–159

    Article  PubMed  CAS  Google Scholar 

  • Wurm F.M, Petropoulos C.J (1994) Plasmid integration, amplification and cytogenetics in CHO cells: questions and comments. Biologicals 22:95–102

    Article  PubMed  CAS  Google Scholar 

  • Yang F, Cai S, Xu L, Feng L (1992) Establishment of hybridoma cell line secreting specific monoclonal antibodies against turnip mosaic virus and analysis of properties of the McAb. Chin J Biotechnol 8:247–254

    PubMed  CAS  Google Scholar 

  • Yoshikawa T, Nakanishi F, Ogura Y, Oi D, Omasa T, Katakura Y, Kishimoto M, Suga KI (2001) Flow cytometry: an improved method for the selection of highly productive gene-amplified CHO cells using flow cytometry. Biotechnol Bioeng 74:435–442

    Article  PubMed  CAS  Google Scholar 

  • Zeyda M, Borth N, Kunert R, Katinger H (1999) Optimization of sorting conditions for the selection of stable, high-producing mammalian cell lines. Biotechnol Prog 15:953–957

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We would like to thank Erika Holroyd, Kathleen Roeder and Angela Davis for laboratory support.

Author information

Authors and Affiliations

  1. Cell Sciences and Development, SAFC Biosciences, 2909 Laclede Ave., St. Louis, MO, 63103, USA

    Jennifer R. Cresswell, Nan Lin, Genova A. Richardson & Kevin J. Kayser

Authors
  1. Jennifer R. Cresswell
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nan Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Genova A. Richardson
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Kevin J. Kayser
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jennifer R. Cresswell .

Editor information

Editors and Affiliations

  1. Dept. Chemical & Biochemical, University College, Dublin, Belfield, Dublin 4, Ireland

    Mohamed Al-Rubeai

Rights and permissions

Reprints and permissions

Copyright information

© 2009 Springer Science+Business Media B.V.

About this chapter

Cite this chapter

Cresswell, J.R., Lin, N., Richardson, G.A., Kayser, K.J. (2009). Cell Xpress™ Applications in Development and Characterization of Biopharmaceutical Recombinant Protein Producing Cell Lines. In: Al-Rubeai, M. (eds) Cell Line Development. Cell Engineering, vol 6. Springer, Dordrecht. https://doi.org/10.1007/978-90-481-2245-5_6

Download citation

Publish with us

Policies and ethics

Search

Navigation