Docetaxel resistance presents a significant obstacle in the treatment of prostate cancer (PCa), resulting in unfavorable patient prognoses. Intratumoral heterogeneity, often associated with epithelial-to-mesenchymal transition (EMT), has previously emerged as a phenomenon facilitating adaptations to various stimuli, thus promoting cancer cell diversity and eventually resistance to chemotherapy, including docetaxel. Hence, comprehending intratumoral heterogeneity is essential for better patient prognosis and the development of personalized treatment strategies. To address this, we employed a high-throughput single-cell flow cytometry approach to identify a specific surface fingerprint associated with docetaxel-resistance in PCa cells and complement it with protein composition analysis of extracellular vesicles We further performed validation of selected antigens using docetaxel-resistant patient-derived xenografts in vivo and determine a 6-molecule surface fingerprint associated with docetaxel resistance in primary PCa specimens. Remarkably, we observed consistent overexpression of CD95 and SSEA-4 surface antigens in both in vitro and in vivo docetaxel-resistant models and in a cell subpopulation of primary PCa tumors exhibiting EMT features. Furthermore, CD95, along with the essential enzymes involved in SSEA-4 synthesis, ST3GAL1, and ST3GAL2, displayed a significant increase in PCa patients undergoing docetaxel-based therapy, correlating with poor survival outcomes. In conclusion, we demonstrate that the identified 6-molecule surface fingerprint associated with docetaxel resistance pre-exists in a subpopulation of primary PCa tumors before docetaxel treatment. This fingerprint, therefore, deserves further validation as a promising tool for predicting docetaxel response in PCa patients before therapy initiation.