Abstract
In 1998, a development occurred in stem cell biology with the first report of the derivation of a human embryonic stem cell (hESC) line. Since then a number of techniques have been used to derive and characterise hESCs. Here, we describe the derivation methods used by our laboratory for isolation of the ICM by immunosurgery and outgrowth of the whole blastocyst. We have added protocols for routine culture, passaging and cryopreservation of our hESC lines as well as the methods we have used for characterisation (flow cytometry, karyotyping, immunocytochemistry, in vitro and in vivo differentiation). Additionally, we have included gene sequences for PCR and an antibody list for immunocytochemistry.
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Acknowledgments
The authors wish to thank patients for donating embryo and staff of the IVF Clinic, Odense University Hospital and Odense IVF for embryo collection and culture. Funding was provided by the Velux Foundation, and from a local grant from the University Hospital of Odense and the local government of Southern Denmark.
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Harkness, L., Kassem, M. (2012). Procedures for Derivation and Characterisation of Human Embryonic Stem Cells from Odense, Denmark. In: Turksen, K. (eds) Human Embryonic Stem Cells Handbook. Methods in Molecular Biology, vol 873. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-794-1_3
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DOI: https://doi.org/10.1007/978-1-61779-794-1_3
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