Characterization and Optimization of Gold Nanoparticle-Based Silver-Enhanced Immunoassays

Silver-enhanced nanoparticle-labeled immunoassays provide a simple, low-cost, and effective way of detecting antigens in dilute solutions. The physical mechanisms behind their operation, however, have not been fully investigated. We present a semiquantitative approach for optimizing sandwich nanoparticle immunoassays using an adsorption-controlled kinetic model. Primary antibodies were immobilized on a solid substrate to bind the target antigens in solution. An optical signal was measured by secondary labeling of antigens with gold nanoparticles and their enhancement by silver nucleation. The opacity of the silver-enhanced spots was quantified by densitometry. The selectivity of the sandwich immunoassays was adequately high, and antigen concentrations as low as 0.1 μg cm-3 (4 ng total) were detected reproducibly. The role of mass transfer was investigated, and a model was developed to optimize the performance of immunoassays by correlating the opacities of silver spots to the concentration and incubation times of antigens and gold nanoparticles. The results could allow the development of more rapid and reliable nanoparticle immunoassays.