Monoclonal antibody-based dual-label time-resolved fluorometric assays in a simplified one-step format

We have performed a one-step all-in-one dual-label dry reagent time-resolved fluorometric assay measuring free and total prostate-specific antigen (PSA). All assay-specific components were predried in an ordinary microtitration well, including the Eu and Tb chelate-labeled monoclonal antibodies for the recognition of free and total PSA, respectively. Only the sample and assay buffer needed to be added. Optimally, equilibrium was reached in <15 min. New intrinsically fluorescent Eu and Tb chelates were used, which made it possible to read the fluorescence directly from the solid phase. The developed dual-label assay with the simple protocol is ideal for easy automation, as the reaction wells can be of alternative design. The performance characteristics are equivalent to those of the present state-of-the-art single-label assays. The detection limits for free and total PSA were 0.03 and 0.02 microg/L, respectively, and the assays were linear over the range tested (up to 300 microg/L).