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Detection and quantitation of circulating tumor cells from solid epithelial tumors could become a valuable tool for therapy monitoring if the procedure can be standardized. In the present work we assessed the influence of preanalytical handling, storage and white blood cell isolation on analysis of a population of spiked tumor cell-line cells and intrinsically present epithelial cells in the peripheral blood of breast and lung cancer patients and the sensitivity of their detection. Sucrose density separation did not enrich epithelial cells, and even depleted them, leading to a gross underestimation of their numbers (3/13 positive, between 2.9 and 50cells/mL) in comparison to red blood cell lysis (13/13 positive, between 77,200 and 800cells/mL). Short-term storage of whole blood samples for up to 7days had little influence on the number of epithelial cells recovered. The effectiveness of magnetic bead enrichment was dependent on the number of relevant cells and the volume used for enrichment. Red blood cell lysis and fluorochrome-labeled antibody staining in a no-wash procedure with subsequent laser scanning cytometry allowed the detection of circulating epithelial cells in 92% of breast and lung cancer patients. Two examples of how this method can be applied for the longitudinal analysis in individual patients are shown, with an increase in numbers preceding relapse and a decrease paralleling tumor reduction. The proposed simple and easy method allows close monitoring, which may help in real-time analysis of the response of solid tumors, especially their systemic component, to therapy and hopefully will contribute to more individually tailored therapy.

Standardized quantification of circulating peripheral tumor cells from lung and breast cancer