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Isolation, cultivation and characterization of adult murine prostate stem cells

Nature Protocols volume 5, pages 702–713 (2010)Cite this article

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Abstract

The successful isolation and cultivation of prostate stem cells will allow us to study their unique biological properties and their application in therapeutic approaches. Here we describe step-by-step procedures on the basis of previous work in our laboratory for the harvesting of primary prostate cells from adolescent male mice by a modified enzymatic procedure; the isolation of an enriched population of prostate stem cells through cell sorting; and the cultivation of prostate stem cells in vitro and characterization of these cells and their stem-like activity, including in vivo tubule regeneration. Normally, it will take approximately 8 h to harvest prostate cells, isolate the stem cell–enriched population and set up the in vitro sphere assay. It will take up to 8 weeks to analyze the unique properties of the stem cells, including their regenerative capacity in vivo.

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Figure 1
Figure 2: Harvest of the urogenital system.
Figure 3: Prostate micro-dissection.
Figure 4: FACS plots of the LSC and LSCT populations.
Figure 5: Representative overview images of spheres, colonies, and prostatic tubules.
Figure 6: Differentiation marker stains of spheres, colonies and prostatic tubules.

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Acknowledgements

We thank Houjian Cai and Yang Zong for the helpful comments on the protocols and for showing the procedure in Supplementary Figure 2. R.U.L. was supported by the California Institute for Regenerative Medicine training grants (T1-00005 and TG2-01169). A.S.G. was supported by Ruth L. Kirschstein National Research Service Award GM07185. This work has also been supported by the Prostate Cancer Foundation Challenge Award for Defining Targets and Biomarkers in Prostate Cancer Stem Cells. O.N.W. is an investigator of the Howard Hughes Medical Institute.

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Author notes

  1. Devon A Lawson

    Present address: Present address: Department of Anatomy, University of California, San Francisco, California, USA.,

Authors and Affiliations

  1. Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, California, USA

    Rita U Lukacs, Devon A Lawson & Owen N Witte

  2. Molecular Biology Institute, University of California, Los Angeles, California, USA

    Andrew S Goldstein

  3. Howard Hughes Medical Institute, University of California, Los Angeles, California, USA

    Donghui Cheng & Owen N Witte

  4. Department of Medical and Molecular Pharmacology, David Geffen School of Medicine, University of California, Los Angeles, California, USA

    Owen N Witte

  5. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, California, USA

    Owen N Witte

Authors
  1. Rita U Lukacs
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  2. Andrew S Goldstein
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  3. Devon A Lawson
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  4. Donghui Cheng
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  5. Owen N Witte
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Contributions

All the authors contributed to the development of the methodology. R.U.L., A.S.G. and O.N.W. contributed to the description of the protocols.

Note: Supplementary information is available via the HTML version of this article.

Corresponding author

Correspondence to Owen N Witte.

Supplementary information

Supplementary Figure 1

Control plots for the FACS sorts (JPG 741 kb)

Supplementary Figure 2

Images of the subrenal prostate regeneration assay (JPG 550 kb)

Supplementary Figure 3

Images of tubules regenerated in the subrenal prostate regeneration assay. (JPG 433 kb)

Supplementary Table 1

Antibodies used for FMO (All Minus One) FACS compensation (see REAGENTS for company and catalogue number information). (PDF 55 kb)

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Lukacs, R., Goldstein, A., Lawson, D. et al. Isolation, cultivation and characterization of adult murine prostate stem cells. Nat Protoc 5, 702–713 (2010). https://doi.org/10.1038/nprot.2010.11

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