Quantification of breast cancer cells in peripheral blood samples by real-time rt-PCR.

BACKGROUND Circulating tumour cells (CTCs) are cells that have detached from a primary tumour, circulate in the peripheral blood, and are considered to be the main root of distant metastases. We present a method for the detection of CTCs by real-time PCR on different cytokeratin markers. MATERIALS AND METHODS Blood samples of a healthy donor were mixed with specific numbers of cells from different breast carcinoma cell line cells. RNA was isolated from the samples and transcribed into cDNA. TaqMan real-time PCR for cytokeratins 8, 18 and 19 was carried out and was correlated to that of 18S. RESULTS Cytokeratin gene expression increased in all samples, when as few as 10 tumour cells were added. In the CAMA-1 cell line, the increase was even greater the more cells were added. CONCLUSION By this methodology, cells from mammary carcinoma cell lines can be detected in blood samples. Its benefit will be validated in samples from patients with breast cancer.