Immunoassay on a single microparticle: the effect of particle size and number on a miniaturized time-resolved fluorometric assay of free prostate-specific antigen Original Research Article

Individual microparticles can be applied to miniaturize conventional immunometric and DNA hybridization assays. In the present report, the effect of particle size and number on the performance of a non-competitive free prostate-specific antigen (PSA) assay were studied in a microliter scale volume. PSA was captured onto the surface of a microparticle by a monoclonal antibody. Particle-bound PSA was detected by time-resolved fluorometry after the release of europium ions from the europium-labeled monoclonal antibody into a fluorescence enhancement solution; 1.8 amol of free PSA was detected in a miniaturized immunoassay by using a single microparticle. The kinetics of the multiple microparticle assay (particle diameter 4 μm, 500 000 particles per assay) was rapid due to a favorable surface-to-volume ratio. In the assay employing a single microparticle (particle diameter 920 μm), the equilibrium was reached in 7.5 min when a total reaction volume of 1 μl was used. This study indicates that a miniaturized assay with a single microparticle as the solid phase is feasible and competitive in terms of kinetics, detection limit and dynamics as an immunoassay format.