title: Murine mammary stem/progenitor cell isolation: Different method matters?
abstract: Murine mammary stem/progenitor cell isolation has been routinely used in many laboratories, yet direct comparison among different methods is lacking. In this study, we compared two frequently used digestion methods and three sets of frequently used surface markers for their efficiency in enriching mammary stem and progenitor cells in two commonly used mouse strains, C57BL/6J and FVB. Our findings revealed that the slow overnight digestion method using gentle collagenase/hyaluronidase could be easily adopted and yielded reliable and consistent results in different batches of animals. In contrast, the different fast digestion protocols, as described in published studies, yielded high percent of non-epithelial cells with very few basal epithelial cells liberated in our hands. The three sets of markers tested in our hands reveal rather equally efficiency in separating luminal and basal cells if same fluorochrome conjugations were used. However, the tendency of non-epithelial cell inclusion in the basal cell gate was highest in samples profiled by CD24/CD29 and lowest in samples profiled by CD49f/EpCAM, this is especially true in mammary cells isolated from C57BL/6J mice. This finding will have significant implication when sorted basal cells are used for subsequent gene expression analysis.
authors.affiliation: School of Laboratory Medicine and Life Science, Wenzhou Medical University, University Town, Wenzhou, 325035 China ; Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Gao
authors.affiliation: School of Laboratory Medicine and Life Science, Wenzhou Medical University, University Town, Wenzhou, 325035 China ; Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA ; Institute of Environmental Safety and Human Health, Wenzhou Medical University, University Town, Wenzhou, 325035 China.
authors.forename: Qiaoxiang
authors.initials: Q
authors.lastname: Dong
authors.affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA ; Institute of Environmental Safety and Human Health, Wenzhou Medical University, University Town, Wenzhou, 325035 China.
authors.forename: Yuanhong
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA ; Institute of Environmental Safety and Human Health, Wenzhou Medical University, University Town, Wenzhou, 325035 China.
authors.forename: Fuchuang
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA ; Institute of Environmental Safety and Human Health, Wenzhou Medical University, University Town, Wenzhou, 325035 China.
authors.forename: Anqi
authors.initials: A
authors.lastname: Wu
authors.affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA ; Institute of Environmental Safety and Human Health, Wenzhou Medical University, University Town, Wenzhou, 325035 China.
authors.forename: Yuanshuo
authors.initials: Y
authors.lastname: Shi
authors.affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA.
authors.forename: Abhik
authors.initials: A
authors.lastname: Bandyopadhyay
authors.affiliation: Flow Cytometry Facility, University of Texas Health Science Center, San Antonio, TX 78299 USA.
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Daniel
authors.affiliation: Institute of Environmental Safety and Human Health, Wenzhou Medical University, University Town, Wenzhou, 325035 China.
authors.forename: Changjiang
authors.initials: C
authors.lastname: Huang
authors.affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78299 USA.
authors.forename: Lu-Zhe
authors.initials: LZ
authors.lastname: Sun
keywords: Digestion method; Enrichment; Flow cytometric analysis; Mammary stem/progenitor cell; Surface marker