Isolation, Propagation, and Clonogenicity of Intestinal Stem Cells

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.