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Isolation, Propagation, and Clonogenicity of Intestinal Stem Cells

  • Prashanthi Ramesh
  • Aleksandar Buryanov Kirov
  • David Johannes Huels
  • Jan Paul MedemaEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2002)

Abstract

Intestinal stem cell research has greatly aided our understanding of the biology of intestinal self-renewal but has also shed light on the role of cancer stem cells (CSCs) in carcinogenesis, cancer growth, and dissemination. With new possibilities for CSC targeting, there is a need to have established techniques for quantifying (cancer) stem cell clonogenicity, particularly in organoid cultures. Here, we describe a detailed methodology for the isolation and expansion of mouse intestinal crypts from three different locations—the colon, proximal, and distal small intestine. In addition, we describe techniques that allow the measurement of stem cell clonogenicity and its manipulation using two approaches—organoid counting and immunohistochemistry.

Keywords

Cancer stem cells Clonogenicity Colorectal cancer Immunohistochemistry Isolation Organoids 

Notes

Acknowledgments

This work was supported by Oncode Institute, Transcan-2 grant Tactic and Dutch Cancer Society (KWF) Grants UvA2015-7587 and 10150. D.J. Huels was supported by an EMBO long-term fellowship ALTF 1102-2017.

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Copyright information

© Springer Science+Business Media New York 2018

Authors and Affiliations

  • Prashanthi Ramesh
    • 1
    • 2
  • Aleksandar Buryanov Kirov
    • 1
    • 2
  • David Johannes Huels
    • 1
    • 2
  • Jan Paul Medema
    • 1
    • 2
    Email author
  1. 1.LEXOR, Center for Experimental and Molecular MedicineAmsterdam UMC, University of Amsterdam, Cancer Center AmsterdamAmsterdamNetherlands
  2. 2.Oncode InstituteAmsterdamNetherlands

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