(A) In normal colon, all epithelial cells, identifiable by their expression of EpCAM, also express CD133. In primary tumors, malignant epithelium is EpCAM⁺CD133⁺, whereas stromal and inflammatory cells are the only EpCAM^–CD133^– cells present, in addition to the extracellular matrix. Thus, the CD133^– cell fraction does not contain cancerous epithelial cells, and therefore, not surprisingly, CD133^– cells do not form tumors in NOD/SCID mice. Only once the disease has progressed to metastasis does a fraction of the EpCAM⁺CD133^– cells appear. During this progression, the precise point during which this subpopulation of EpCAM⁺CD133^– cells emerges is unknown. However, in their study in this issue of the JCI, Shmelkov et al. (5) demonstrate in liver metastases that both CD133⁺ and CD133^– populations of EpCAM⁺ colon cancer epithelial cells are capable of forming tumors in NOD/SCID mice. Interestingly, the CD133^– tumors grow more aggressively than the CD133⁺ tumors. (B) Shmelkov et al. (5) suggest that the angle and location of colon tissue sections may confound the analysis of CD133 expression in vivo. CD133 protein is localized to the apical surface of the epithelial cells. Depending on the orientation of crypts in the section, CD133 expression could be underestimated if the luminal surface of cells is not exposed, resulting in false-negative staining for CD133.