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Derivation of a new human embryonic stem cell line, Endeavour-2, and its characterization

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Abstract

Here, we describe the derivation of a novel human embryonic stem cell (hESC) line, Endeavour-2 (E-2), propagated on human fetal fibroblasts (HFF) in a serum-replacement media. The inner cell mass (ICM) was manually dissected from the blastocyst without using immunodissection and, therefore, antibodies from animal sources. A total of 20 embryos were thawed and cultured, eight embryos were hatched, and five ICMs were obtained. They were transferred onto HFF used as feeder layer, and one colony representing the initial cell proliferation of a new hESC line, E-2, was obtained. The newly emerged hESC colony was passaged first by physical dissection and subsequently by enzymatic dissociation. E-2 has been in culture for over 6 months and has been shown to possess typical features of a pluripotent hESC line including expression of stem cell surface markers (SSEA4, TRA-160, and integrin alpha-6), intracellular alkaline phosphatase, and pluripotency gene markers, OCT4 and NANOG. This hESC line shows lineage-specific differentiation into various representative cell types expressing markers characteristic of the three somatic germ layers under both in vitro and in vivo conditions. E-2 line shows a normal karyotype (46 XX) and has been successfully cryopreserved and thawed several times using slow-freezing procedures. E-2 adds to the repertoire of existing hESC lines for research and development purposes in the field of regenerative medicine.

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Acknowledgments

This study was supported by the Sydney Foundation for Medical Research, Diabetes Australia Research Trust and the Australian Foundation for Diabetes Research. We thank Bo Yuan and Jaemin Kim for their help in tissue culture, Leonie Gaudry for assistance with FACS (SEALS, Prince of Wales Hospital), and Pauline Dalzell Pauline Dalzell, (Cytogenetics Laboratory, SEALS Prince of Wales Hospital) for assistance with karyotyping and Emeritus Professor Doug Saunders from IVF Australia for organizing egg donations.

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Authors and Affiliations

  1. Diabetes Transplant Unit, Prince of Wales Hospital & University of New South Wales, Sydney, NSW, Australia

    Kuldip S. Sidhu, Justin G. Lees & Bernard E. Tuch

  2. Stem Cell Lab, School of Biotechnology and Biomolecular Sciences and School of Psychiatry, Faculty of Medicine, University of New South Wales, 408 Wallace Wurth Building, Sydney, NSW, Australia

    Kuldip S. Sidhu

  3. IVF Australia, Sydney, NSW, Australia

    John P. Ryan

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  1. Kuldip S. Sidhu
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Correspondence to Kuldip S. Sidhu.

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Editor: J. Denry Sato

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Sidhu, K.S., Ryan, J.P., Lees, J.G. et al. Derivation of a new human embryonic stem cell line, Endeavour-2, and its characterization. In Vitro Cell.Dev.Biol.-Animal 46, 269–275 (2010). https://doi.org/10.1007/s11626-010-9280-8

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  • DOI: https://doi.org/10.1007/s11626-010-9280-8

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