Nanoencapsulated microcrystalline particles for superamplified biochemical assays

We report on the preparation and utilization of a novel class of particulate labels based on nanoencapsulated organic microcrystals with the potential to create highly amplified biochemical assays. Labels were constructed by encapsulating microcrystalline fluorescein diacetate (FDA; average size of 500 nm) within ultrathin polyelectrolyte layers of poly(allylamine hydrochloride) and poly(sodium 4-styrenesulfonate) via the layer-by-layer technique. Subsequently, the polyelectrolyte coating was used as an "interface" for the attachment of anti-mouse antibodies through adsorption. A high molar ratio of fluorescent molecules present in the microcrystal core to biomolecules on the particle surface was achieved. The applicability of the microcrystal-based label system was demonstrated in a model sandwich immunoassay for mouse immunoglobulin G detection. Following the immunoreaction, the FDA core was dissolved by exposure to organic solvent, leading to the release of the FDA molecules into the surrounding medium. Amplification rates of 70-2000-fold (expressed as an increase in assay sensitivity) of the microcrystal label-based assay compared with the corresponding immunoassay performed with direct fluorescently labeled antibodies are reported. Our approach provides a general and facile means to prepare a novel class of biochemical assay labeling systems. The technology has the potential to compete with enzyme-based labels as it does not require long incubation times, thus speeding up bioaffinity tests.