Abstract
Two different approaches have been adopted for the cryopreservation of human embryonic stem cells (hESCs): vitrification and conventional slow cooling/rapid warming. The vitrification method described here is designed for hESCs that grow as discrete colonies on a feeder cell monolayer, and are subcultured by manual subdivision of the colonies into multicellular clumps. hESCs that are subcultured by enzymatic dissociation can more conveniently be cryopreserved by conventional slow cooling/rapid warming methods. Although both methods are suitable for use in a research context, neither is suitable for cryopreservation of embryonic stem cells destined for clinical diagnostic or therapeutic uses without modification.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Thomson, J. A., Itskovitz-Eldor, J., Shapiro, S. S., et al. (1998) Embryonic stem cell lines derived from human blastocysts. Science 282, 1145–1147.
Hoffman, L. M. and Carpenter, M. K. (2005) Characterisation and culture of human embryonic stem cells. Nature Biotech. 23, 699–708.
Robertson, E. J. (1987) Embryo derived stem cell lines. In: Teratocarcinomas and Embryonic Stem Cells: A Practical Approach, (Robertson, E. J., ed.), IRL Press, Oxford, UK, pp. 71–112.
Chan, S. Y. and Evans, M. J. (1991) In situ freezing of embryonic stem cells in multiwell plates. Trends Genetics 7, 76.
Ure, J. M., Fiering, S., and Smith, A. G. (1992) A rapid and efficient method for freezing and recovering clones of embryonic stem cells. Trends Genetics 8, 6.
Heng, B. C., Kuleshova, L. L., Bested, S. M., Liu, H., and Cao, T. (2005) The cryopreservation of human embryonic stem cells. Biotechnol. Appl. Biochem. 41, 97–104.
Amit, M., Carpenter, M. K., Inokuma, M. S., et al. (2000) Clonally derived human embryonic stem cell lines maintain pluripotency. Dev. Biol. 227, 271–278.
Reubinoff, B. E., Pera, M. R, Fong, C. Y., Trounson, A., and Bongso, A. (2000) Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro. Nature Biotech. 18, 399–405.
Reubinoff, B. E., Pera, M. E, Vajta, G., and Trounson, A. (2001) Effective cryopreservation of human embryonic stem cells by the open pulled straw vitrification method. Human Reprod. 16, 2187–2194.
Richards, M. K., Fong, C.-Y., Tan, S., Chan, W.-K., and Bongso, A. (2004) An efficient and safe xeno-free cryopreservation method for the storage of human embryonic stem cells. Stem Cells 22, 779–789.
Zhou, C. Q., Mai, Q. Y., Li, T., and Zhaung, G. J. (2004) Cryopreservation of human embryonic stem cells by vitrification. Chin. Med. J. (Engl). 117, 1050–1055.
Ji, L., de Pablo, J. J., and Palecek, S. P. (2004) Cryopreservation of adherent human embryonic stem cells. Biotechnol. Bioeng. 88, 299–312.
Cowan, C. C., Klimanskaya, I., McMahon, J., et al. (2004) Derivation of embryonic stem-cell lines from human blastocysts. N. Engl. J. Med. 350, 1353–1356.
Ware, C. B., Nelson, A. M., and Blau, C. A. (2005) Controlled-rate freezing of human ES cells. BioTechniques 38, 879–883.
Yang, P. E, Tsung, T. C., Cheng, Q. K., Wu, C. E, and Cao, Y. L. (2005) Effective cryopreservation of human embryonic stem cells by programmed freezing. Shi Yan Sheng Wu Xue Bao 38, 247–256.
Ha, S. Y., Jee, B. C., Suh, C. S., et al. (2005) Cryopreservation of human embryonic stem cells without the use of a programmable freezer. Human Reprod. 20, 1779–1785.
Vajta, G., Holm, P., Kuwayama, M., et al. (1998) Open pulled straw (OPS) vitrification: a new way to reduce cryoinjuries of bovine ova and embryos. Mol. Rep. Dev. 51, 53–58.
ESI Methodology Manual Embryonic Stem Cell Culture, July 2005. (http://www.escellinternational.com). Last accessed June 6, 2006.
Heng, B. C., Bested, S. M., Chan, S. H., and Cao, T. (2005) A proposed design for the cryopreservation of intact and adherent human embryonic stem cell colonies. In Vitro Cell. Dev. Biol.-Animal 41, 77–79.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2007 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Hunt, C.J., Timmons, P.M. (2007). Cryopreservation of Human Embryonic Stem Cell Lines. In: Day, J.G., Stacey, G.N. (eds) Cryopreservation and Freeze-Drying Protocols. Methods in Molecular Biology™, vol 368. Humana Press. https://doi.org/10.1007/978-1-59745-362-2_18
Download citation
DOI: https://doi.org/10.1007/978-1-59745-362-2_18
Publisher Name: Humana Press
Print ISBN: 978-1-58829-377-0
Online ISBN: 978-1-59745-362-2
eBook Packages: Springer Protocols