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Generation and Establishment of Murine Adherent Cell Lines

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Basic Cell Culture Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 946))

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Abstract

We describe a method to derive cell lines and clones from cells of the murine midgestation aorta-gonads-mesonephros (AGM) microenvironment. We start from subdissected AGM regions in “explant” or “single cell suspension” type cultures from embryos transgenic for tsA58, a temperature-sensitive mutant of the SV40 T antigen gene. The number of cells in such cultures initially expand, but in most cases, this expansion phase is followed by a stable or even decline in cell number. After this so-called crisis phase, cell proliferation is noticeable in more than 90% of the cultures. Stromal cell clones can be isolated from these cultures, some of which have been cultured for more than 50 population doublings, and functionally characterized using various methods These stromal cell clones are valuable tools for the study of the regulation of hematopoietic stem and progenitor cells in the midgestation mouse embryo.

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Acknowledgement

We gratefully acknowledge Jessyca Maltman (Terry Fox Laboratory, BC Cancer Agency, Vancouver, BC, Canada) for her help and experience in the culture of adherent mouse marrow cells. We also would like to thank Elaine Dzierzak for the opportunity to establish cell lines from midgestation embryos.

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Authors and Affiliations

  1. The Stem Cell Physiology Laboratory, Medizinische Klinik, Technische Universität München, Munich, Germany

    Rouzanna Istvanffy & Robert A. J. Oostendorp

Authors
  1. Rouzanna Istvanffy
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  2. Robert A. J. Oostendorp
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Corresponding author

Correspondence to Robert A. J. Oostendorp .

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Editors and Affiliations

  1. , Experimental Therapeutics, BC Cancer Agency, West 10th Ave. 675, Vancouver, V5Z 1L3, British Columbia, Canada

    Cheryl D. Helgason

  2. STEMCELL Technologies, Inc., W. Seventh Avenue 570, Vancouver, V5Z 1B3, British Columbia, Canada

    Cindy L. Miller

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Istvanffy, R., Oostendorp, R.A.J. (2013). Generation and Establishment of Murine Adherent Cell Lines. In: Helgason, C., Miller, C. (eds) Basic Cell Culture Protocols. Methods in Molecular Biology, vol 946. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-128-8_19

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  • DOI: https://doi.org/10.1007/978-1-62703-128-8_19

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-127-1

  • Online ISBN: 978-1-62703-128-8

  • eBook Packages: Springer Protocols

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