3D culture assays of murine mammary branching morphogenesis and epithelial invasion

Kim Vy Nguyen-Ngoc, Eliah R. Shamir, Robert J. Huebner, Jennifer N. Beck, Kevin J. Cheung, Andrew J. Ewald

Research output: Chapter in Book/Report/Conference proceedingChapter

69 Scopus citations


Epithelia are fundamental tissues that line cavities, glands, and outer body surfaces. We use three-dimensional (3D) embedded culture of primary murine mammary epithelial ducts, called "organoids," to recapitulate in days in culture epithelial programs that occur over weeks deep within the body. Modulating the composition of the extracellular matrix (ECM) allows us to model cell-and tissue-level behaviors observed in normal development, such as branching morphogenesis, and in cancer, such as invasion and dissemination. Here, we describe a collection of protocols for 3D culture of mammary organoids in different ECMs and for immunofluorescence staining of 3D culture samples and mammary gland tissue sections. We illustrate expected phenotypic outcomes of each assay and provide troubleshooting tips for commonly encountered technical problems.

Original languageEnglish (US)
Title of host publicationTissue Morphogenesis
Subtitle of host publicationMethods and Protocols
PublisherSpringer New York
Number of pages28
ISBN (Electronic)9781493911646
ISBN (Print)9781493911639
StatePublished - Sep 22 2014


  • 3D culture
  • Branching morphogenesis
  • Collagen I
  • Extracellular matrix
  • Immunofluorescence
  • Mammary epithelium
  • Matrigel
  • Organoids
  • Organotypic culture

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)


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