Phenotypic and Genetic Characterization of Circulating Tumor Cells by Combining Immunomagnetic Selection and FICTION Techniques

Abstract

S U M M A R Y The presence of circulating tumor cells (CTCs) in breast cancer patients has been proven to have clinical relevance. Cytogenetic characterization of these cells could have crucial relevance for targeted cancer therapies. We developed a method that combines an immunomagnetic selection of CTCs from peripheral blood with the fluorescence immunophenotyping and interphase cytogenetics as a tool for investigation of neoplasm (FICTION) technique. Briefly, peripheral blood (10 ml) from healthy donors was spiked with a predetermined number of human breast cancer cells. Nucleated cells were separated by double density gradient centrifugation of blood samples. Tumor cells (TCs) were immunomagnetically isolated with an anti-cytokeratin antibody and placed onto slides for FICTION analysis. For immunophenotyping and genetic characterization of TCs, a mixture of primary monoclonal anti-pancytokeratin antibodies was used, followed by fluorescent secondary antibodies, and finally hybridized with a TOP2A/HER-2/CEP17 multicolor probe. Our results show that TCs can be efficiently isolated from peripheral blood and characterized by FICTION. Because genetic amplification of TOP2A and ErbB2 (HER-2) in breast cancer correlates with response to anthracyclines and herceptin therapies, respectively, this novel methodology could be useful for a better classification of patients according to the genetic alterations of CTCs and for the application of targeted therapies. (J Histochem Cytochem 56:667-675, 2008)

Screening and Molecular Analysis of Single Circulating Tumor Cells Using Micromagnet Array

“…The control cell, which is a leukocyte here, shows the ratio number to be 1. Observation of SK-BR-3 (HER-2 positive) and MDA-MB-231 (HER-2 negative) cells here accords well with the previous conclusions 40 41 . It approves the efficacy of our system in providing retrievable cells for molecular studies.…”

Characterization of circulating tumor cells by fluorescence in situ hybridization

“…Drawback of this finding is that assessment of the genetic makeup of the tumor becomes less informative as fewer CTC are available and therapeutic targets can thus be assessed in fewer patients. Recent reports using alternative technologies suggest the presence of a significant larger number of CTC in carcinoma patients this however may well be contributed to the definition of what constitutes a CTC (39–46). Still, one would like to assess the presence or absence of therapeutic targets in as large a patient group as possible urging the need for technology that increase the yield of CTC.…”

Detection of micrometastases in peripheral blood of non-small cell lung cancer with a refined immunomagnetic nanoparticle enrichment assay

“…Immunomagnetic nanoparticle enrichment combined with ICC and pathology examination provides a new method for early diagnosis of NSCLC. 14,15 In recent years, the continuous development of nanotechnology has seen the successful development of iron nanomaterials. We used nanopure iron microspheres as the core of immunomagnetic nanoparticles.…”