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C. Mazzini, P. Pinzani, D. Massi, F. Salvianti, F. Ucci, U. Menchini, M. Pazzagli; Biomolecular Characterization of Circulating Tumour Cells (CTC) by "ISET" (Isolation by Size of Epithelial Tumor Cells) in Uveal Melanoma Patients: A New Approach to the Study of CTC. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5954.
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The aim of our study was the counting and the morphological characterization of circulating tumor cells (CTCs) by the Isolation by Size of Epithelial Tumor cells (ISET) method in the peripheral blood of uveal melanoma patients. An evaluation of the method’s ability to reveal the presence of occult tumor cells in uveal melanoma patient blood was performed and the results compared to quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay for the evaluation of Tyrosinase mRNA expression.
ISET is a technique to directly isolate CTCs. It is based on filtration of peripheral blood through polycarbonate membrane with pores of calibrated 8 µm diameter. Basing on pores dimension, the filter can thus retain larger cells (i.e. cancerous cells), but not leukocytes whose diameter is usually smaller than the pores. The major advantages of the ISET is that the method is very simple and fast, consisting in only one filtration step, while other system for CTCs separation require multiple stages with the risk of loosing rare cells. It allows to exactly count the number of cells per ml of blood: the filter is suited for the isolation of both single CTC and groups of CTCs (microemboli) which seems to posses a higher metastatic potential. Cells on the filter are fixed and intact so that they can be analyzed by immunological, cytologycal or biomolecular techniques. Furthermore, cells can be recovered from the filter by laser assisted microdissection and further characterized by molecular analysis. Finally, the method is very sensitive, being capable of detecting a single tumour cell added to 1 ml of blood . ISET was carried out using a device for filtration provided by Metagenex (Paris, France), a disposable module of filtration, and a polycarbonate membrane
We find a correlation between the two methods used to detect CTC. ISET is always positive in metastatic patients.
The comparison between ISET (direct method) and Real Time RT-PCR (indirect method ) performed in the same patient and in the same sample is a unique opportunity of our study as no studies are available in melanoma patients using this approach. This comparison will give important insights on the methodological practicability and clinical validity of these two different approaches for CTC detection.Authors disclose any Commercial Relationship.
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