Expression of EpCAM in human breast cancer cell lines and primary epithelial cells. (A) Protein expression was analyzed by Western Blot analysis with an antibody directed against the extracellular domain of EpCAM. Primary cells (HMECs) showed weak or no expression of EpCAM. Epithelial tumor cells displayed strong EpCAM expression, whereas EpCAM expression decreased in more mesenchymal tumor cells. The two different bands represent glycosylated and basic isoforms of EpCAM. Tubulin served as internal loading control. (B) Densitometric analysis of EpCAM protein expression of two independent experiments. Values indicate relative intensity units. (C) EpCAM gene expression was analyzed by real-time PCR analysis using TATA-Box binding protein and GAPDH as internal housekeeping genes. All cell lines were analyzed in triplicate and normalized to the expression levels of MCF-10A cells. (D) Tumor cell lines were phenotyped by immunofluorescence analysis after staining for epithelial markers cytokeratin-18 and E-cadherin and the mesenchymal marker vimentin. Magnification 400x.