Abstract Background This study evaluated the diagnostic performance of the Atellica® IM High-Sensitivity PSA (hsPSA)* assay in men >50 years undergoing prostate cancer (PC) screening in conjunction with digital rectal exam (DRE). Methods Serum samples were collected prospectively from 2671 subjects >50 years referred to a urologist for evaluation of PC from 26 sites across the United States. To establish PC versus benign disease, all subjects underwent a transrectal prostate biopsy ≤60 days after blood draw. At the cutoff of 4.0 ng/mL for the hsPSA assay, the percentage of PCs detected (sensitivity) and corresponding percentage of biopsies avoided in men without PC (specificity) were determined. Results Cancer prevalence (malignant biopsy) was 58.5% (1562/2671) among the diagnostic population. Preliminary results demonstrated that for DRE alone, sensitivity and specificity were 30.9% (482/1562) and 83.0% (920/1109) compared to biopsy. For hsPSA alone at the cutoff of 4.0 ng/mL, sensitivity was 88.7% (1386/1562). The table provides incidence of cancer and positive biopsies by subgroups defined by DRE and hsPSA. hsPSA detected 61.1% (955/1562) of PCs that DRE did not. Of those DRE+, 71.8% (482/671) had positive biopsy. Of those DRE+ and hsPSA ≥4.0 ng/mL, 78.6% (431/548) had positive biopsy, which significantly increased PC detection by 7.0% (95% CI:1.4%-12.5%). Similarly, in the DRE- population, while 54.0% of subjects had positive biopsy, subjects both DRE- and hsPSA <4.0 ng/mL had only 38.5% positive biopsies, a significant difference of 16.5% (95% CI: 6.3%-27.8%). Conclusions Using DRE alone detected only 3 out of 10 patients with PC. It was observed that when used in conjunction with DRE, the hsPSA assay improved both sensitivity and specificity of PC detection over DRE alone. CAUTION: Investigational Device. Not available for sale. Future availability cannot be guaranteed. The results are preliminary and were achieved in unique setting with no expected timeline for completion.