title: Limiting-dilution transplantation assays in mammary stem cell studies.
abstract: Mammary reconstitution assays can be used to measure the stem cell frequency within an epithelial population by transplanting increasingly diluted single-cell preparations of the population of interest. There are fundamental steps in the single-cell isolation protocol which are directly related to the number of single epithelial cells obtained. Once single-cell suspensions have been obtained, serial dilutions are prepared and transplanted into the cleared fat pads of the host mice. After 8-10 weeks, the transplanted fat pads are reevaluated for the presence of epithelial outgrowths. Based on the frequency of no outgrowth for each one of the transplanted dilutions, it is possible to estimate the frequency of mammary repopulating cells present in a given cell population. Here, we give details on how to carry out all these steps.
authors.affiliation: Langone School of Medicine, New York University, New York, NY 10016, USA.
authors.forename: Irineu
authors.initials: I
authors.lastname: Illa-Bochaca
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Fernandez-Gonzalez
authors.forename: Dawne N
authors.initials: DN
authors.lastname: Shelton
authors.forename: Bryan E
authors.initials: BE
authors.lastname: Welm
authors.forename: Carlos
authors.initials: C
authors.lastname: Ortiz-de-Solorzano
authors.forename: Mary Helen
authors.initials: MH
authors.lastname: Barcellos-Hoff
mesh_terms: D000273:Adipose Tissue / Q000166:cytology / Q000637:transplantation; D000818:Animals; D049109:Cell Proliferation; D002469:Cell Separation; D017690:Cell Transplantation / Q000379:methods*; D002498:Centrifugation; D004847:Epithelial Cells / Q000637:transplantation; D008321:Mammary Glands, Animal / Q000166:cytology*; D051379:Mice; D013194:Staining and Labeling; D013234:Stem Cells / Q000166:cytology*