• cancer stem cells;
  • breast cancer;
  • flow cytometry;
  • immunohistochemistry;
  • immunofluorescence;
  • cytokeratin;
  • CD44;
  • CD90;
  • CD133;
  • CD117



A variety of markers have been proposed to identify breast cancer stem cells. Here, we used immunohistostaining and flow cytometry to analyze their interrelationships and to sort cells for tumorigenicity studies.


Cytokeratin, CD44, and CD90 were localized to primary breast cancer and normal breast (NB) tissue by immunohistostaining and related to CD117 and CD133 expression by flow cytometry. Immunodeficient NOD.CB17-Prkdcscid/J and NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice were used to test tumorigenicity of sorted CD90+ low-light scatter, CD90+ high-light scatter, and CD90neg tumor cells.


NB basal cells coexpressed CD44 and CD90. As cells transited luminally, CD44 was retained and downmodulated, and CD90 was lost and cytokeratin increased. In breast tumors, basal-like CD44+/CD90+ cells were localized to the tumor periphery, adjacent to CD90+ stroma. Like normal luminal cells, interior tumor cells were CD44+/CD90−. Immunophenotyping (CD44/CD90/CD117/CD133) of cytokeratin+ cells revealed no significant difference in expression between tumors and tumor-free breast. In both, CD133 was distributed approximately equally among CD44/CD90 subsets, whereas CD117 expression was highest in the basal-associated CD44+/CD90+ subset. Sorted CD90+ pleural effusion cells with lymphoid light scatter, 49% of which were CD44+, were uniquely tumorigenic in immunodeficient mice (100 cells/injection).


Our data demonstrate that all tumors contain a small population of CD44+/CD90+ cells, mimicking the phenotype of ductal-basal cells. These are localized to the tumor periphery, adjacent to CD90+ stroma. Among the nonhematopoietic, nonmesothelial cells found in metastatic pleural effusions, low-light scatter CD90+ cells are most potently tumorigenic, compared to high-scatter CD90+ cells and CD90− cells. © 2010 International Clinical Cytometry Society