Human breast cell lines are enriched for basal and mesenchymal phenotypes. (a) Normal breast cell lines demonstrate loss of EpCAM+/CD49f- and EpCAM+/CD24+/CD49f+ populations compared to primary breast epithelial cells isolated from reduction mammoplasty. Reduction mammoplasty tissues (RM) and normal breast cells lines as well as matched RM with HME cell lines were stained with EpCAM, CD24 and CD49f and quantified by flow cytometry as described in Materials and methods. (b) Quantification of mammospheres formed in non-adherent culture by HME II, MCF10A and MCF10F cell lines (left) and representative images (right). Bar = 100 μm. (c, d) Addition of 5% serum to the culture conditions of HME cells increases differentiation to a more luminal state as assessed by flow cytometry for EpCAM, CD49f and CD24. Representative dot plots are shown in C and quantification in D. (e) Changes in Basal/Luminal differentiation were assessed by immunofluorescence staining for CK8/18, EpCAM, CK14 and vimentin following treatment with serum. Cells were counterstained with DAPI (blue). HME I cells are shown, bar = 100 μm.