Magnetic-particle-based, ultrasensitive chemiluminescence enzyme immunoassay for free prostate-specific antigen

• A low-cost and rapid assay for f-PSA in serum was developed using MMP-based CLEIA.
• f-PSA detection: 0.1–30 ng mL−1 concentration range; 0.1 ng mL−1 detection limit.
• The proposed method showed high sensitivity, good reproducibility and stability.
• The strategy showed great potential in the fabrication of MMP-based f-PSA test kits.

We report a magnetic-particle (MMP)-based chemiluminescence enzyme immunoassay (CLEIA) for free prostate-specific antigen (f-PSA) in human serum. In this method, the f-PSA is sandwiched between the anti-PSA antibody coated MMPs and alkaline phosphatase (ALP)-labeled anti-f-PSA antibody. The signal produced by the emitted photons from the chemiluminescent substrate (4-methoxy-4-(3-phosphatephenyl)-spiro-(1,2-dioxetane-3,2′-adamantane)) is directly proportional to the amount of f-PSA in a sample. The present MMP-based assay can detect f-PSA in the range of 0.1–30 ng mL−1 with the detection limit of 0.1 ng mL−1. The linear detection range could match the concentration range within the “diagnostic gray zone” of serum f-PSA levels (4–10 ng mL−1). The detection limit was sufficient for measuring clinically relevant f-PSA levels (>4 ng mL−1). Furthermore, the method was highly selective; it was unaffected by cross-reaction with human glandular kallikrein-2, a kallikrein-like serine protease that is 80% similar to f-PSA. The proposed method was finally applied to determine f-PSA in 40 samples of human sera. Results obtained using the method showed high correlation with those obtained using a commercially available microplate CLEIA kit (correlation coefficient, 0.9821). This strategy shows great potential application in the fabrication of diagnostic kits for determining f-PSA in serum.

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