| Summary: | <p>Abstract</p> <p>Background</p> <p>Immunomagnetic enrichment followed by RT-PCR (immunobead RT-PCR) is an efficient methodology to identify disseminated carcinoma cells in the blood and bone marrow. The RT-PCR assays must be both specific for the tumor cells and sufficiently sensitive to enable detection of single tumor cells. We have developed a method to test RT-PCR assays for any cancer. This has been investigated using a panel of RT-PCR markers suitable for the detection of breast cancer cells.</p> <p>Methods</p> <p>In the assay, a single cell line-derived tumor cell is added to 100 peripheral blood mononuclear cells (PBMNCs) after which mRNA is isolated and reverse transcribed for RT-PCR analysis. PBMNCs without added tumor cells are used as specificity controls. The previously studied markers epidermal growth factor receptor (<it>EGFR</it>), mammaglobin 1 (<it>MGB1</it>), epithelial cell adhesion molecule (<it>EpCAM</it>/<it>TACSTD1</it>), mucin 1 (<it>MUC1</it>), carcinoembryonic antigen (<it>CEA</it>) were tested. Two new epithelial-specific markers <it>ELF3</it> and <it>EphB4</it> were also tested.</p> <p>Results</p> <p><it>MUC1</it> was unsuitable as strong amplification was detected in 100 cell PBMNC controls. Expression of <it>ELF3</it>, <it>EphB4</it>, <it>EpCAM</it>, <it>EGFR</it>, <it>CEA</it> and <it>MGB1</it> was found to be both specific for the tumor cell, as demonstrated by the absence of a signal in most 100 cell PBMNC controls, and sensitive enough to detect a single tumor cell in 100 PBMNCs using a single round of RT-PCR.</p> <p>Conclusions</p> <p><it>ELF3</it>, <it>EphB4</it>, <it>EpCAM</it>, <it>EGFR</it>, <it>CEA</it> and <it>MGB1</it> are appropriate RT-PCR markers for use in a marker panel to detect disseminated breast cancer cells after immunomagnetic enrichment.</p>
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