title: Quantifying Epithelial Early Common Progenitors from Long-Term Primary or Cell Line Sphere Culture.
abstract: Here, a protocol to quantify epithelial early common progenitor/stem cells grown as spheres in non-adherent culture conditions is described. This protocol is based on the combination of two functional tests: the sphere assay to maintain and enrich early progenitor/stem cells, and the epithelial colony-forming cells (E-CFC) assay to identify and quantify further differentiated epithelial progenitors. Primary spheres mainly contain progenitors and rare stem/early common progenitor cells while secondary and tertiary spheres contain progenitor cells derived from the early common progenitor/stem cell population maintained through passages and partially differentiated. Spheres are enzymatically and mechanically dissociated; the derived cells are subsequently plated on irradiated NIH-3T3 fibroblasts for further processing, as in the E-CFC assay. The principle of this assay is to quantify the number of epithelial colonies generated by cells present in the different sequential spheres. This assay has therefore been named the early common progenitor-derived colonies assay (ECP-DC).
authors.affiliation: INSERM U1052, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|Université de Lyon, Université Lyon 1, Lyon, France.
authors.forename: Flora
authors.initials: F
authors.lastname: Clément
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Renji-Med X Clinical Stem Cell Research Center, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.|School of Biomedical Engineering & Med-X Research Institute, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Helen He
authors.initials: HH
authors.lastname: Zhu
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Renji-Med X Clinical Stem Cell Research Center, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.|School of Biomedical Engineering & Med-X Research Institute, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Wei-Qiang
authors.initials: WQ
authors.lastname: Gao
authors.affiliation: INSERM U1052, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|Université de Lyon, Université Lyon 1, Lyon, France.|Department of Tumor Escape Signaling, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|Centre Léon Bérard, Lyon, France.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Delay
authors.affiliation: INSERM U1052, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Lyon, France.|Université de Lyon, Université Lyon 1, Lyon, France.|Department of Tumor Escape Signaling, Centre de Recherche en Cancérologie de Lyon, Lyon, France.
authors.forename: Véronique
authors.initials: V
authors.lastname: Maguer-Satta
mesh_terms: D000818:Animals; D001940:Breast / Q000166:cytology; D001943:Breast Neoplasms / Q000473:pathology; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002460:Cell Line; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D018920:Coculture Techniques / Q000379:methods; D004847:Epithelial Cells / Q000166:cytology*; D005260:Female; D006801:Humans; D008297:Male; D051379:Mice; D041681:NIH 3T3 Cells; D011467:Prostate / Q000166:cytology; D018874:Spheroids, Cellular / Q000166:cytology*; D013234:Stem Cells / Q000166:cytology*
keywords: cell lines; epithelial colonies; human stem cells; in vitro quantification; spheres