An ultrasensitive immunoassay for prostate-specific antigen based on conventional colorimetric detection

Objective: Development of an ultrasensitive immunoassay for serum PSA involving conventional detection probes. Design and Methods: The assay involves a polyclonal antibody immobilized in microtitration Wells and a monoclonal antibody labeled with horseradish peroxidase. In a one-step assay, the enzymatic activity of the bound detection antibody is monitored by the addition of hydrogen peroxide/tetramethylbenzidine substrate reagent followed by spectrophotometric quantification of the conversion product. Results: The assay has a lower detection limit of 0.003 μg/L, biological detection limit of 0.009 μg/L, and intra- and inter-assay CVs of 8.2% and 10.5% at PSA concentrations of 0.022 and 0.065 μg/L, respectively. The recovery of the assay averaged 104% and it demonstrated a dilution linearity down to at least 0.01 μg of PSA/L. Results of comparison data correlated well with those obtained by a well established enzyme immunoassay. The serum PSA concentrations were <0.012 μg/L in the majority of patients (53.8%) who had undergone radical prostatectomy. Conclusions: This assay is well suited for post-surgical monitoring of PSA in patients with prostate cancer.