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The characterization of circulating tumor cells (CTCs) can lead to a promising strategy for monitoring residual or relapsing prostate cancer (PCa) after local therapy. The aim of this study was to compare three innovative technologies for CTC enumeration in 131 high-risk patients with PCa, before and after radiotherapy, combined with androgen deprivation. The CTC number was tested using the FDA-cleared CellSearch® system, the dual fluoro-EPISPOT assay that only detects functional CTCs, and the in vivo CellCollector® technology. The highest percentage of CTC-positive patients was detected with the CellCollector® (48%) and dual fluoro-EPISPOT (42%) assays, while the CellSearch® system presented the lowest rate (14%). Although the concordance among methods was only 23%, the cumulative positivity rate was 79%. A matched-pair analysis of the samples before, and after, treatment suggested a trend toward a decrease in CTC count after treatment with all methods. CTC tended to be positivity correlated with age for the fluoro-EPISPOT assay and with PSA level from the data of three assays. Combining different CTC assays improved CTC detection rates in patients with non-metastatic high-risk PCa before and after treatment. Our findings do not support the hypothesis that radiotherapy leads to cancer cell release in the circulation. [ABSTRACT FROM AUTHOR]