We comparatively evaluated different cytokeratin (CK) reagents analyzed by flow cytometry (FCM) for the identification of the best combination of DNA/CK staining for detecting minimal numbers of breast cancer cells in peripheral blood (PB). In 59 primary breast cancer tumors, we comparatively analyzed the reactivity for up to 6 different anti-CK reagents using multiparameter FCM: anti-CK7, anti-CK20, anti–pan-CK, anti-CK8/CK18, anti-CK8, and anti-CK18. Afterward, dilutional experiments of Michigan Cancer Foundation (MCF)7 breast cancer cells in PB were performed, and the sensitivity of a DNA/CK18 staining was evaluated.

Our results showed that anti-CK18 reagents were those providing the brightest and more sensitive staining for primary breast cancer tumor cells by FCM. Dilutional experiments of MCF cells in PB showed that the DNA/anti-CK18 double staining was highly specific for the identification of epithelial cells; its sensitivity ranged between 10–6 and 10–7 (detection of 1 tumor cell among 106 to 107 nucleated blood cells). Combined assessment of DNA cell contents and reactivity for CK18 by FCM is a sensitive method for the specific identification of breast cancer cells in PB.

Author notes

Both authors contributed equally.
Supported by grant PI021078 from the Fondo de Investigaciones Sanitarias (FIS), Ministerio de Sanidad y Consumo, Madrid, Spain.