Microtextured substrates and microparticles used as in situ lenses for on-chip immunofluorescence amplification.

We propose a concept for enhancing the fluorescent detection signal of on-chip immunoassays by using three-dimensional 3 and 8 µm size microtextured structures as substrates for the assay, which allows exploitation of a large number of fluorophores within the focal plane of an optical microscope for detection. Additionally, we demonstrate the use of 3 and 9.75 µm dielectric microparticles, respectively, as in situ lenses on the 3 and 8 µm microstructures, respectively, for further enhancement of the optical signal. In our model system, the fluorescent complex is formed on (3-aminopropyl)triethoxysilane microstructures and we use carboxyl-functionalized melamine microparticles as the in situ lenses. Mouse IgG diluted in phosphate-buffered saline is used as model target antigen and can be easily detected down to a concentration of 2 ng/mL thanks to these approaches. Also, we present a detailed two-dimensional numerical study of the light propagation through a dielectric microparticle using the finite element method, providing key insight into the signal amplification mechanism of a microlens, and point out its advantageous use in microfluidic assays.