Clonal MaCSC differentiation. A. Clonal single cells from FVB/N cell line Py9813 are K8K14-positive. Left panel stained with secondary antibody alone. B. Early colonies contain only double-positive cells. Scale bar 20 μm. C. In two-dimensional cultures, K8 + K14+ Py9813 cells can self-renew or differentiate to form single-positive K8 or K14-positive cells. Scale bar 20 μm. D. Py9813 cells form hollow mammospheres that show the presence of double-positive cells and K8 single-positive luminal cells. Single-positive myoepithelial cells are less commonly observed. Scale bar 20 μm. E. The percentage of double-positive, single-positive and double-negative cells in immunofluorescence-stained two-dimensional cultures. Analysis of variance showed the groups were highly significantly different, P <0.0001 and by Holm-Sidak’s multiple comparisons test, there were significantly fewer K8+ and K14+ cells compared with K8 + K14+ cells, P <0.0001, and significantly fewer K14+ cells than K8+ cells, P <0.01 in both cell lines. No double-negative cells were observed in cultures of either cell type. Data are means ± SEM. F. Cultures of control or 300 nM retinoic acid-treated Py9813 cells stained with oil red O. Scale bar 20 μm. (PDF 900 kb
