Probing quenched dye fluorescence of Cy3-DNA-Au-nanoparticle hybrid conjugates using solution and array platforms

Tuning the luminescence intensity of fluorophores using nanoparticles has shown great potential for the detection of inorganic metal ions, viruses, and proteins. The enhancement or quenching of a dye's fluorescence intensity is strongly dependent on the spatial separation of the dye from the nanoparticle surface. To extend luminescence probing from the solution platform to the solid-state platform, we explored and performed dye quenching assessment using an array format in this study. We report the distance-dependent fluorescence behavior of Au-DNA conjugates prepared by equilibrating phosphine-stabilized gold nanoparticles (AuNPs) of 10-nm size with the designed spacer ds-DNA consisting of thiol-modified target and Cy3-labeled complementary probe of different lengths (5-20 nm). The Cy3-labeled products were immobilized onto MPTMS (3-mercaptopropyltrimethoxysilane)-modified glass substrates and then excited with a 532-nm laser source. Quenching efficiency of AuNPs with increasing Au-to-dye distance was assessed using ligand exchange of the thiolated oligonucleotide by 2-mercaptoethanol (ME) to obtain free Cy3-DNA probe, thus eliminating nanoparticle effect on the dye's luminescence intensity. Effective exchange, revealed by UV-vis absorption and fluorescence profiles, was achieved in a few minutes. It was observed that fluorescence quenching of Au-DNA-Cy3 assessed using the array format was consistent with the result in solution phase for the conjugates with up to 10-nm Au-to-Cy3 separation distance.