Immunoassay for the discrimination of free prostate-specific antigen (fPSA) forms with internal cleavages at Lys145 or Lys146 from fPSA without internal cleavages at Lys145 or Lys146

Abstract

Total levels of circulating prostate-specific antigen (tPSA) are strongly associated with prostate cancer (PCa) risk and outcome but benign prostate disease is the most frequent cause of a moderately elevated PSA level. Free PSA (fPSA) forms are independently associated with PCa risk and contribute modest diagnostic enhancements above and beyond tPSA alone. We developed an immunoassay for fPSA subfractions containing internal cleavages at Lys145 or Lys146 (fPSA-N). The assay was based on blocking intact single-chain fPSA (fPSA-I) with antibody 4D4 which does not detect PSA containing internal cleavages at Lys145 or Lys146. We also measured fPSA-N in blood from healthy volunteers and in anti-coagulated plasma from 76 men with or without evidence of PCa at biopsy. The analytical and functional detection limits of this assay were 0.016 ng/mL and 0.10 ng/mL, respectively. The median recovery of male fPSA-N from female plasma was 95.0 %. All 12 female samples (average age 28 years) had fPSA-N concentrations at or below the analytical detection limit. The median fPSA-N concentration (0.050 ng/mL) in 9 healthy male volunteers (age < 40 years) was below the functional detection limit, 0.420 ng/mL in 27 patients with benign prostate conditions and 0.239 ng/mL in 49 patients with PCa. Deming regression analysis of the patient samples showed that the measured fPSA-N concentrations were generally 23% lower than the previously calculated (fPSA minus fPSA-I) concentrations, likely due to differences in the antibody combinations used. In conclusion, we have developed a sensitive, specific and direct immunoassay for fPSA-N which can be used to study the clinical relevance of this PSA isoform.

A Four-kallikrein Panel and β-Microseminoprotein in Predicting High-grade Prostate Cancer on Biopsy: An Independent Replication from the Finnish Section of the European Randomized Study of Screening for Prostate Cancer

“…Cryopreserved sample aliquots (serum or plasma) were shipped to the Wallenberg Research Laboratories at Lund University (Malmö, Sweden) for assay of the four kallikrein markers and MSP during 2014-2015. The analyses for total and free PSA [18], intact PSA [19], hK2 [20], and MSP [21] were previously reported [22,23]. [26].…”

Detection of High Grade Prostate Cancer among PLCO Participants Using a Prespecified 4-Kallikrein Marker Panel

“…Immunoassay measurements of total and free PSA, 15 intact PSA, 16 hK2, 17 and MSP, 18 were conducted on AutoDelfia ® 1235 automatic immunoassay systems (Wallenberg Research Laboratories, Department of Translational Medicine, Lund University, Skåne University Hospital, Malmö, Sweden). Free and total PSA were measured using the dual-label DELFIA Prostatus ® total/free PSA-Assay (Perkin-Elmer, Turku, Finland) calibrated against the World Health Organization (WHO) 96/670 (PSA-WHO) and WHO 68/668 (free PSA-WHO) standards.…”

Can one blood draw replace transrectal ultrasonography‐estimated prostate volume to predict prostate cancer risk?

“…To study the independent role of nicked PSA in the present model, a direct immunoassay for nicked PSA is needed instead of the calculated value. A previously published nicked PSA immunoassay needs improvements to suit routine clinical use .…”