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

Individual microparticles can be applied to miniaturize conventional immuno metric and DNA hybridization assays. In the present report, the effect of p article size and number on the performance of a non-competitive free prosta te-specific antigen (PSA) assay were studied in a microliter scale volume. PSA was captured onto the surface of a microparticle by a monoclonal antibo dy. 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 mu m, 500 000 partic les per assay) was rapid due to a favorable surface-to-volume ratio. In the assay employing a single microparticle (particle diameter 920 mu m), the e quilibrium was reached in 7.5 min when a total reaction volume of 1 mu l wa s used. This study indicates that a miniaturized assay with a single microp article as the solid phase is feasible and competitive in terms of kinetics , detection limit and dynamics as an immunoassay format. (C) 1999 Elsevier Science B.V. All rights reserved.