Multilayers of hydrogels loaded with microparticles: a fast and simple approach for microarray manufacturing.

We present a novel and simple approach towards the creation of arrays of biomolecules for the multiplexed detection of biological interactions. Microarrays were obtained by cutting stacked layers of biofunctionalized polystyrene particle layers embedded in a permeable agarose matrix. Microparticles were therefore the vehicles for biorecognition. The three-dimensional constructs were obtained by consecutive dipping steps in a pre-gel solution. Our strategy enables the rapid manufacturing of a large number of array copies in a flexible manner and without any specialized instrumentation. Model binding assays for the detection of rabbit and mouse IgG were performed as a proof of concept using a fluorescence microscope for read-out. The limits of detection were in the low picomolar range for the sandwich assay while 1 IgG out of 50,000 background proteins could be detected in a reverse phase assay. Thus, without any assay optimization, sensitivities comparable to the ones usually observed for standard fluorescence-based assays were achieved with the particle/hydrogel array.