Prostate adenocarcinoma often presents at a late stage, due to a lack of early clinical symptoms and lack of accurate objective markers. This study aimed to identify and validate proteomics-based biomarkers useful for prostate cancer diagnosis and to establish a marker-panel for prostate cancer and benign prostate hyperplasia (BPH). Global protein expression patterns in fresh tissue specimens from 8 patients with prostate carcinoma and 16 with BPH were analyzed by two-dimensional gel electrophoresis. Differentially expressed proteins were identified by MALDI-TOF mass spectrometry. We compared our results with those of published studies and defined a set of common biomarkers. We identified 22 differentially expressed proteins between BPH and prostate carcinomas. The up-regulated proteins in cancer compared to BPH included protein disulfide-isomerase, 14-3-3-protein, Enoyl CoA-hydrase, prohibitin and B-tubulin β-2. Keratin-II, desmin, HSP71, ATP-synthase-β-chain and creatine kinase-β-chain were down-regulated. Survey of the literature showed that 15 of our 22 identified proteins have been previously reported to differ in their expression levels between BPH and prostate cancer by other laboratories. The expression patterns of these biomarkers could successfully cluster BPH and adenocarcinomas as well as prostate cancer of low and high Gleason scores. This study validates protein-biomarkers that can be useful for accurate diagnosis and prognostic monitoring of prostate adenocarcinoma. Despite varied prevalence of the disease between different ethnic populations (i.e., high in Sweden, low in Saudi Arabia); the biomarkers indicate that BPH and prostate cancers are biologically 'homogeneous' in their protein expression patterns across wide geographical regions.