Cackowski FC, Wang Y, Decker JT, et al. - Experts intended to create novel strategies to determine and, if discovered, investigate localized prostate cancer (PCa) patient disseminated tumor cells (DTCs). The putative DTC population was present in ten of 58 patients with localized PCa, four of eight patients with metastatic PCa of varying disease control, and one of eight patients with no known cancer, at a threshold of four cells per million BM cells, and was positively associated with patients’ plasma prostate-specific antigen values. Heightened expression of PCa marker genes in four of eight patients with localized PCa was exhibited by RNA-Seq analysis of the putative DTC population collected from samples above (three patients) and below (five patients) the threshold of 4 putative DTCs per million, however, not the one normal donor who had the putative DTC population present. In the gene features of PCa, in two of three localized PCa patients, whole-exome sequencing also demonstrated the presence of single nucleotide polymorphisms and structural variants. To investigate the likely contaminating cell types, a myeloid colony formation assay, differential counts of cell smears, and analysis of the RNA-Seq data using the CIBERSORT algorithm, which most strongly suggested the presence of B-cell lineages as a contaminant, were used. Lastly, EPCAM enrichment and polymerase chain reaction for PCa markers to determine DTC prevalence was used and it discovered evidence of DTCs in 21 of 44 samples. Thus, in the BM of a subset of patients with localized PCa, the presence of DTCs and report a novel fluorescence-activated cell sorting method for isolation and analysis of viable DTCs was supported.