Introduction: Liquid biopsy, especially when performed by the isolation, expansion, and examination of circulating tumor cells (CTCs) from peripheral blood, has become an innovative and transforming diagnostic tool in Clinical Oncology. The CTCs have already entered the clinical practice as an alternative method to invasive tumor biopsy for detecting postsurgical and/or posttreatment minimal residual disease, to predict cancer recurrence and real-time treatment response. In this context, the retrospective observational project, known as CHARACTEX, has permitted to state that it is possible to exploit blood-based cytologic samples through short-term culture and in vitro CTC expansion. Methods: This method is based initially on a gradient-sedimentation technique, which impoverishes without completely depriving the obtained sample from the hematological cells, followed by short-term (14 days) in vitro culture and expansion and cytomorphological and flow cytometric analysis to investigate whether the expanded cell population possesses proliferative advantage and fits with criteria, which are consistent to the known primary tumor. Results: The originality of this method is that, apart from the above exposed criteria, there is no selection bias for the isolation of the cells from peripheral blood (like immunomagnetic bead treatment or preliminary immunocytochemistry), which can potentially introduce some limitation to the cell population under evaluation. Conclusion: The examination of the expanded cell population obtained by this method is very rewarding for both the pathologist – who can assess multiple tumor-related variables (like immunocytochemistry, flow cytometry of several parameters, and molecular pathology on cell suspensions and cell blocks obtained from them) – and the clinician.

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