CD133 has been associated with cell properties such as self renewal, migration and vasculogenic mimicry, potentially involved in generation of circulating tumor cells (CTCs). We characterized CD133 expression in CTCs of 98 no-metastatic breast cancer patients. CTCs were isolated by immunomagnetic techniques using magnetic beads labelled with a multi-cytokeratin(CK)-specific antibody (CK3-11D5) and CTCs and CD133 detection through immunocytochemical methods. CK(+) /CD133(+) CTCs were identified in 65% of patients at baseline and 47,8% after systemic therapy (p=0.53). Correlation of CD133 status in CTCs with classical clinicopathological characteristics and response to therapy was performed. Her2 not amplified and low Ki-67 index were positively correlated with presence of CK(+) /CD133(+) CTCs. Before any treatment, CK(+) /CD133(+) CTCs were more frequently isolated in patients with luminal breast cancer subtype. No statistically significant differences were found between proportion of CK(+) /CD133(+) CTCs and BC subtypes after systemic therapy, implying a relative enrichment of CK(+) /CD133(+) CTCs in triple negative and HER2-amplified tumors. While we CK(+) /CTCs decreases after chemotherapy when analyzing the whole population, CK(+) /CD133(+) CTCs were enriched in post-treatment samples in non-luminal BC subtypes suggesting the potential role of CD133 as a promising marker of chemoresistance in these patient. Further prospective studies and extensive pre-clinical modeling will be needed to confirm whether CD133 is a marker of resistance to chemotherapy, and its role as a target for novel anticancer therapies targeting cancer stem cells and tumor vasculature. © 2013 Wiley Periodicals, Inc.