CD44+/CD24-/low/ESA+ cells in human breast cancer cell lines enrich for tumorigenicity. Shown are representative flow cytometry analysis and gates for epithelial-specific antigen (ESA), CD24 and CD44 sorting. (a) SUM159 cells were sorted on ESA. Dot plots show resulting CD44+/CD24-/low/ESA+ or CD44+/CD24-/ESA- fractions. (b) SUM149 cells were sorted on CD44 and CD24 and subsequently sorted for ESA+, shown in histograms. (c) Histological analysis of SUM159 and SUM149 orthotopic tumors. The upper panels present representative hematoxylin and eosin stained sections at 20× magnification of tumors that formed from 5 × 105 unsorted, 1,000 CD44+/CD24-/low/ESA+, or 1 × 106 CD44+/CD24-/ESA- SUM159 cells. Note that the tumor histology from CD44+/CD24-/ESA- sorted cells is different than that from unsorted or CD44+/CD24-/low/ESA+ cells. The lower panels present hematoxylin and eosin stained sections at 20× magnification of tumors that formed from 1 × 105 unsorted, CD44+/CD24-/low/ESA+, or CD44+/CD24+/ESA+ sorted SUM149 cells. Note the tumor histology from CD44+/CD24+/ESA+ sorted cells is markedly different from that of the unsorted or the CD44+/CD24-/low/ESA+ sorted cells.