Current Developments in Cell Culture Technology

  • Glyn Stacey
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 745)


The ideal features of a cell culture system for in vitro investigation depend on what questions the system is to address. However, in general, highly valuable systems will replicate the characteristics and more specifically, the responses, of normal human tissues. Systems that can faithfully replicate different tissue types provide tremendous potential value for in vitro research and have been the subject of much research effort in this area over many years. Furthermore, a range of such systems that could mimic key genetic variations or diseases would have special value for toxicology and drug discovery. In the pursuit of such model systems, there are a number of significant practical issues to consider for their application, which includes ability to deliver with ease, the required quantities of cells at the time needed. In addition any cell culture assay will need to be robust and reliable and provide readily interpreted and quantified endpoints. Other general criteria for cell culture systems include scalability to provide the very large cell numbers that may be required for high throughput systems, with a high degree of reliability and reproducibility. The amenability of the cell culture for down-scaling may also be important, to permit the use of very small test samples (e.g., in 96-well arrays), even down to the level of single cell analysis. This chapter explores the range of new cell culture systems for scaling up cell cultures that will be needed for high throughput toxicology and drug discovery assays. It also reviews the increasing range of novel systems that enable high content analysis from small cell numbers or even single cells. The hopes and challenges for the use of human stem cell lines are also investigated in comparison with the range of eukaryotic cells types currently in use in toxicology.


Cell Culture System Stem Cell Line hESC Line Roller Bottle Rotating Wall Vessel 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    MacLeod RAF, Dirks WG, Matsuo Y et al. Widespread intra-species cross-contamination of human tumour cell lines arising at source. Int J Cancer 1999; 83:555–563.PubMedCrossRefGoogle Scholar
  2. 2.
    Masters JR, Thomson, JA, Daly-Burns B et al. Short tandem repeat profiling provides an international reference standard for human cell lines. Proc Natl Acad Sci USA 2001; 98:8012–8017.PubMedCrossRefGoogle Scholar
  3. 3.
    Rottem S, Naot Y. Subversion and exploitation of host cells by mycoplasma. Trends in Microbiol 1998; 6:436–440.CrossRefGoogle Scholar
  4. 4.
    Stacey GN. Sourcing human embryonic stem cell lines. In: Sullivan S, Cowan C. and Eggan K, eds. Human Embryonic Stem Cells: A Practical Handbook. Chichester: John Wiley and Sons, 2007:11–24.CrossRefGoogle Scholar
  5. 5.
    Stacey GN, Hartung T. Availability, standardisation and safety of human cells and tissues for drug screening and testing. In: Marx U, Sandig V, eds. Drug Testing In Vitro: Breakthroughs and Trends in Cell Culture Technology. Weinheim: Wiley-VCH Verlag, 2006:231–246.Google Scholar
  6. 6.
    Robitski AA, Rothermerl A. An overview on bioelectronic and biosensoric microstructures supporting high-content screening in cell cultures. In: Marx U, Sandig V, eds. Drug Testing In Vitro: Breakthroughs and Trends in Cell Culture Technology. Weinheim: Wiley-VCH Verlag, 2006:79–97.CrossRefGoogle Scholar
  7. 7.
    Dickson JA, Suzangar M. A predictive in vitro assay for the sensitivity of human solid tumours to hyperthermia (42 degrees C) and its value in patient management. Clin Oncol 1976; 2:141–55PubMedGoogle Scholar
  8. 8.
    Chambard M, Verrier B, Gabrion J et al. Polarization of thyroid cells in culture: evidence for the basolateral localization of the iodide “pump” and of the thyroid-stimulating hormone receptor-adenyl cyclase complex. J Cell Biol 1983; 96:1172–1177.PubMedCrossRefGoogle Scholar
  9. 9.
    Elvin P, Wong V, Evans CW. A study of the adhesive, locomotory and invasive behaviour of Walker 256 carcinosarcoma cells. Exp Cell Biol 1985; 53:9–18.PubMedGoogle Scholar
  10. 10.
    McCall E, Povey J, Dumonde DC. The culture of vascular endothelial cells to confluence on microporous membranes. Thromb Res 1981; 24:417–31.PubMedCrossRefGoogle Scholar
  11. 11.
    Lazarovici P, Li M, Perets A et al. Intelligent biomatrices and engineered tissue constructs: in vitro models for drug discovery and toxicity Testing. In: Marx U, Sandig V, eds. Drug Testing In Vitro: Breakthroughs and Trends in Cell Culture Technology. Weinheim: Wiley-VCH Verlag, 2006:1–51.CrossRefGoogle Scholar
  12. 12.
    McKinney VZ, Rinker KD, Truskey GA. Normal and shear stresses influence the spatial distribution of intracellular adhesion molecule-1 expression in human umbilical vein endothelial cells exposed to sudden expansion flow. J Biomech 2006; 39:806–817.PubMedCrossRefGoogle Scholar
  13. 13.
    Martin Y, Vermette P. Bioreactors for tissue mass culture: design, characterization and recent advances. Biomaterials 2005; 26:7481–7503.PubMedCrossRefGoogle Scholar
  14. 14.
    Kofidis T, Lenz A, Buoblik J et al. Pulsatile perfusion and cardiomyocyte viability in a solid three-dimensional matrix. Biomaterials 2003; 24:5009–5014.PubMedCrossRefGoogle Scholar
  15. 15.
    Klaus DM. Clinostats and bioreactors. Gravit Space Biol Bull 2001; 14:55–64.PubMedGoogle Scholar
  16. 16.
    Bruce MP, Boyd V, Duch C et al. Dialysis-based bioreactor systems for the production of monoclonal Antibodies—alternatives to ascites production in mice. J Immunol Methods 2001; 264:59–68.CrossRefGoogle Scholar
  17. 17.
    Pannell R, Milstein C. An oscillating bubble chamber for laboratory scale production of monoclonal antibodies as an alternative to ascitic tumours. J Immunol Methods 1992; 146:43–48.PubMedCrossRefGoogle Scholar
  18. 18.
    Davis J. Systems for cell culture scale-up. In: Stacey GN, Davis J, eds. Medicines from Animal Cells. Chichester: John Wiley and Sons, 2007:1435–172.Google Scholar
  19. 19.
    Portner R, Geise C. An overview of bioreactor design, prototyping and process control for reproducible three-dimensional tissue culture. In: Marx U, Sandig V, eds. Drug Testing In Vitro: Breakthroughs and Trends in Cell Culture Technology. Weinheim: Wiley-VCH Verlag, 2006:53–70.CrossRefGoogle Scholar
  20. 20.
    Jayme D. Development and optimization of serum-free and protein-free media. In: Stacey GN, Davis J, eds. Medicines from Animal Cells. Chichester: John Wiley and Sons, 2007:29–44.CrossRefGoogle Scholar
  21. 21.
    Coecke S, Balls M, Bowe G et al. Guidance on good cell culture practice. A report of the second ECVAM task force on good cell culture practice. Altern Lab Anim—ATLA 2005; 33(1):1–27.Google Scholar
  22. 22.
    Freshney IR. Culture of Animal Cells: A Manual of Basic Technique, 4th ed. Chichester: John Wiley and Sons, 2004.Google Scholar
  23. 23.
    Greene JJ, Brophy CI. Induction of protein disulfide isomerase during proliferation arrest and differentiation of SH5Y neuroblastoma cells. Cell Mol Biol 1995; 41:473–80.PubMedGoogle Scholar
  24. 24.
    Fleck RA, Athwal H, Bygraves J et al. Optimisation of NB-4 and HL-60 differentiation for use in opsonophagocytosis assays. In Vitro Cell Dev Biol Anim 2003; 39:235–242.PubMedCrossRefGoogle Scholar
  25. 25.
    Howard D, Buttery LD, Shakesheff KM et al. Tissue engineering: strategies, stem cells and scaffolds. J Anat 2008; 213:61–72.CrossRefGoogle Scholar
  26. 26.
    Carswell KS, Weiss JW, Papoutsakis ET. Low oxygen tension enhances the stimulation and proliferation of human T-lymphocytes in the presence of IL-2. Cytotherapy 2000; 2:25–37.PubMedCrossRefGoogle Scholar
  27. 27.
    D’Ippolito G, Diabira S, Howard GA et al. Low oxygen tension inhibits osteogenic differentiation and enhances stemness of human MIAMI cells. Bone 2006; 39:513–22.PubMedCrossRefGoogle Scholar
  28. 28.
    Ezashi T, Das P, Roberts RM. Low O2 tensions and the prevention of differentiation of hESCs. Proc Natl Acad Sci USA 2005; 102:4783–4788.PubMedCrossRefGoogle Scholar
  29. 29.
    Enninga IC, Groenendijk RT, van Zeeland AA et al. Use of low temperature for growth arrest and synchronization of human diploid fibroblasts. Mutat Res 1984; 130:343–352.PubMedGoogle Scholar
  30. 30.
    Fischer SM, Viaje A, Harris KL et al. Improved conditions for murine epidermal cell culture. In Vitro 1980; 16:180–188.PubMedCrossRefGoogle Scholar
  31. 31.
    Yang QR, Berghe DV. Effect of temperature on in vitro proliferative activity of human umbilical vein endothelial cells. Experientia 1995; 51:126–132.PubMedCrossRefGoogle Scholar
  32. 32.
    Takahashi K, Tereda S, Ueda H et al. Growth rate suppression of cultured mammalian cells enhances protein productivity. Cytotechnology 1994; 15:57–64.PubMedCrossRefGoogle Scholar
  33. 33.
    Milstein, J, Grachev V, Padilla A et al. WHO activities towards the three Rs in the development and control of biological medicines. Dev Biol Stand 1996; 86:31–39.Google Scholar
  34. 34.
    Hartung T. Three Rs potential in the development and quality control of pharmaceuticals. ALTEX 2001; 18:3–11.PubMedGoogle Scholar
  35. 35.
    Rubenstein AL. Zebrafish assays for drug toxicity screening. Expert Opin Drug Met Toxicol 2006; 2:231–240.CrossRefGoogle Scholar
  36. 36.
    Scholtz G, Pohl I, Genschow E et al. Embryotoxicity screening using embryonic stem cells in vitro: correlations with in vivo teratogenicity. Cells Tissues Organs 1999; 165:203–211.CrossRefGoogle Scholar
  37. 37.
    Rolletscheck A, Blyszczuk P, Wobus AM. Embryonic stem cell derived cardiac neuronal and pancreatic cells as model systems to study toxicological effects. Physiol Rev 2005; 85:635–67.CrossRefGoogle Scholar
  38. 38.
    Thomson J, Itskovitz-Eldor J, Shapiro S et al, Embryonic stem cell lines derived from human blastocysts. Science 1998; 282:1145–1147.PubMedCrossRefGoogle Scholar
  39. 39.
    Sullivan S, Cowan CA, Eggan K, eds.. Human Embryonic Stem Cells: The Practical Handbook. Chichester: John Wiley and Sons, 2007.Google Scholar
  40. 40.
    Freshney IR, Stacey GN, Auerbach J, eds. Culture of Human Stem Cells. Hoboken: John Wiley and Sons, 2007.Google Scholar
  41. 41.
    Loring JF, Wesselschmidt RL, Schwartz PH, eds. Human Stem Cell Manual, A Laboratory Guide. New York: Academic Press, 2007.Google Scholar
  42. 42.
    Masters JR, Palsson BO, Thomson JA, eds. Human Cell Culture V6, Embryonic Stem Cells. Dordrecht: Springer, 2007.Google Scholar
  43. 43.
    Chojnacki A, Weiss S. Production of neurons, astrocytes and oligodendrocytes from mammalian CNS stem cells. Nat Protoc 2008; 3:935–40.PubMedCrossRefGoogle Scholar
  44. 44.
    Turksen K. Human Embryonic Stem Cell Protocols. Methods Mol Biol 2006; 331:1–12.PubMedGoogle Scholar
  45. 45.
    Roche E, Ensenat-Waser R, Vicente-Salar N et al. Insulin-producing cells from embryonic stem cells experimental considerations. Methods Mol Biol 2007; 407:295–309.PubMedCrossRefGoogle Scholar
  46. 46.
    Pickering SJ, Minger S, Patel M et al. Generation of human embryonic stem cell line encoding the cystic fibrosis mutation deltaF508, using preimplantation genetic diagnosis. Reprod Biomed Online 2003; 10:390–397.CrossRefGoogle Scholar
  47. 47.
    Chen Y, He ZX, Liu A et al. Embryonic stem cells generated by nuclear transfer of human somatic nuclei into rabbit oocytes. Cell Res 2003; 13:251–63.PubMedCrossRefGoogle Scholar
  48. 48.
    Takahashi K, Tanabe K, Ohnuki M et al;. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 2007; 131:861–872.PubMedCrossRefGoogle Scholar
  49. 49.
    Yu J, Vodyanik MA, Smuga-Otto K et al. Induced pluripotent stem cell lines derived from human somatic cells. Science 2007; 318:1917–1920.PubMedCrossRefGoogle Scholar
  50. 50.
    Nakagawa M, Koyanagi M, Tanabe K et al. Generation of induced pluripotent stem cells without Myc from mouse and human fibroblasts. Nat Biotechnol 2008; 26:101–106.PubMedCrossRefGoogle Scholar
  51. 51.
    Jeffs SA. Expression of recombinant biomedical products from continuous mammalian cell lines. In: Stacey GN, Davis J, eds. Medicines from Animal Cells. Chichester: John Wiley and Sons, 2007: 61–78.Google Scholar
  52. 52.
    Chapman EJ, Hurst CD, Pitt E et al. Expression of hTERT immortalises normal human urothelial cells without inactivation of the p16/Rb pathway. Oncogene 2006; 25:5037–5045.PubMedCrossRefGoogle Scholar
  53. 53.
    Morales CP, Gandia KG, Ramirez RD et al. Characterisation of telomerase immortalised normal human oesophageal squamous cells. Gut 2003; 52:327–333.PubMedCrossRefGoogle Scholar
  54. 54.
    Maeda T, Tashiro H, Katabuchi H et al. Establishment of an immortalised human ovarian surface epithelial cell line without chromosomal instability. Br J Cancer 2005; 93:116–123.PubMedCrossRefGoogle Scholar
  55. 55.
    International Stem Cell Banking Initiative. Consensus guidance for banking and supply of human embryonic stem cell lines for research purposes. Stem Cell Rev Rep 2009; 5:301–314.CrossRefGoogle Scholar
  56. 56.
    International Stem Cell Initiative, Adewumi O, Aflatoonian B, Ahrlund-Richter L et al. Characterization of human embryonic stem cell lines by the International Stem Cell Initiative. Nat Biotechnol 2007; 25:803–816.PubMedCrossRefGoogle Scholar
  57. 57.
    Spielmann H. Ethical environment and scientific rationale towards in vitro alternatives to animal testing: where are we going. In: Marx U, Sandig V, eds. Drug Testing In Vitro: Breakthroughs and Trends in Cell Culture Technology. Weinheim: Wiley-VCH Verlag, 2006:251–267.CrossRefGoogle Scholar
  58. 58.
    Anderson R, O’Hare M, Balls M et al. The availability of human tissue for biomedical research—the report and recommendations of ECVAM workshop. Altern Lab Anim—ATLA 1998; 26(6):763–777.Google Scholar

Copyright information

© Landes Bioscience and Springer Science+Business Media 2012

Authors and Affiliations

  • Glyn Stacey
    • 1
  1. 1.Division of Cell Biology and ImagingNational Institute for Biological Standards and ControlSouth Mimms, Potters Bar, HertfordshireUK

Personalised recommendations