Embedded tumor cell calibrators (ETC) for normalized quantitative multi-gene expression analysis of micrometastatic cells in blood and bone marrow of cancer patients

1065 Background: The purpose of this study was the validation of a new preanalytical enrichment and molecular detection method using embedded tumor cell calibrators (ETC) for quantitative gene expression analysis of circulating tumor cells (CTC) in breast cancer patients. Methods: Samples from every patient were devided in native probes and matched calibrator probes spiked with either 2, 4 or 8 breast carcinoma tumor cells. The high affinity antibodies BM7 (MUC-1) and VU1D9 (EpCAM) were used for immunomagnetic tumor cell enrichment of 10 mL peripheral EDTA-blood of patients with primary breast cancer and metastatic disease. Separated cells were used for mRNA isolation and c-DNA synthesis. A real-time quantitative RT-PCR approach for the markers cytokeratin 19 (CK19), survivin (Sur), mammaglobin 1 (MG1) and prostate-specific ets factor (PSE), HER-2, CD276 and FGFR2 was established using primers and FAM-labeled TaqMan probes selected with the Universal-Probe-Library system (Roche AG, Basel, CH). Results: Po...