Abstract
Establishment of robust xenograft models is critical to studying human hematopoiesis in a physiologic setting. Using a recently developed immunodeficient mouse strain, we have established long-term multilineage human grafts and demonstrated their serially transplantability using limited numbers of purified human hematopoietic stem cells (HSCs). Herein, we describe our protocol for the isolation of human HSC (Lin-CD34+CD38−CD90+) from umbilical cord blood (CB) as well as the xenotransplantation system that allows stable engraftment of human hematopoietic cells with as few as ten HSCs. Isolation of CB mononuclear cells requires 2–3 h, and cells may be cryopreserved before transplantation. Isolation of HSC requires approximately 2–3 h, and transplantation requires 1 h. Short-term and long-term engraftment is assessed 4–6 weeks and 10–12 weeks post-transplantation, respectively, with preparation and analysis time requiring 4–8 h at each time point.
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Acknowledgements
We thank Libuse Jerabek and Theresa Storm for their lab management expertise and Adriane Mosley for animal husbandry. This research is supported by National Institutes of Health/National Cancer Institute grants R01CA86017 to I.L.W. and KO8CA1295470 to C.Y.P. R.M. is supported by the Walter and Idun Y. Berry Foundation.
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C.Y.P. and R.M. contributed equally to this work.
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C.Y.P and R.M. have no conflicts of interest to disclose. I.L.W. was a member of the scientific advisory board of Amgen and owns significant Amgen stock. I.L.W. cofounded and consulted for Systemix, is a cofounder and director of Stem Cells Inc., and cofounded Cellerant, Inc.
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Park, C., Majeti, R. & Weissman, I. In vivo evaluation of human hematopoiesis through xenotransplantation of purified hematopoietic stem cells from umbilical cord blood. Nat Protoc 3, 1932–1940 (2008). https://doi.org/10.1038/nprot.2008.194
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DOI: https://doi.org/10.1038/nprot.2008.194
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