Purpose: We previously found a statistically significant number of cytokeratin 19 (CK19)؉ cells in peripheral blood (PB) of stage IV breast cancer (BC) patients compared with those of healthy volunteers, using a quantitative real-time reverse transcription-PCR. We aimed to apply the technique on bone marrow (BM) of primary operable BC patients. Pre-and postoperative PB samples of these patients were further analyzed to investigate possible shedding of CK19؉ cells during the operation.Experimental Design: In 54 primary operable BC patients, we analyzed 50 BM samples taken preoperatively and 297 PB samples. PB samples were collected before surgery; immediately after surgery; on the first, second, and fifth day postoperatively; and one month postoperatively.Results: In BM of controls and BC patients, we detected a median of 28 and 568 CK19؉ cells/5 ؋ 10 6 leukocytes, respectively (P < 0.001). In preoperative blood (B-1) samples, we measured a median of 109 CK19؉ cells. Using the upper limit of 95% confidence interval of controls as cutoff, 74% and 52% of BM and (B-1), respectively were considered CK19؉. There was no significant correlation between CK19؉ cells in BM and (B-1) and classical prognostic factors. We found no significant difference between blood samples at different time points with respect to the average CK19؉ cells. Conclusions:In primary BC patients, we detected high numbers of CK19؉ cells in BM and PB (B-1) samples compared with controls. However, no significant correlation between the presence of CK19؉ cells in BM and PB and classical prognostic factors was found. We detected no statistically significant influence of surgical manipulation on CK19؉ cells.

Plasmid dilutions are more reliable than cell line dilutions for quantification of gene expression, and more objective criteria for positivity could be defined based on the characteristics of the standard curve (slope and intercept). A more universally accepted agreement on the definition of the cut-off value for positivity is needed.

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