DNA detection assay based on fluorescence quenching of rhodamine B by gold nanoparticles: The optical mechanisms

- The inner filter effect and RB quenching by GNPs are basic mechanisms of the fluorescent DNA assay.
- Complete fluorescence quenching correlates with complete adsorption of all RB molecules on GNPs.
- The low detection limit (LDL) of the fluorescent assay is about 30-100 pM of target DNA.
- The fluorescent LDL is close to that for colorimetric and dynamic light scattering DNA assays.

The different ability of single- and double-stranded oligonucleotides to stabilize gold nanoparticles (GNPs) in solution has recently been used to design several label-free hybridization assays on the basis of optical changes associated with GNP aggregation. DNA hybridization can be detected through changes in dye fluorescence quenching by GNPs. Here we examine the mechanisms behind a fluorescent DNA assay for model systems containing DNA oligonucleotides, 15-nm GNPs, and Rhodamine B (RB). There was a direct correlation between complete disappearance of fluorescence and complete adsorption of all RB molecules on nonaggregated GNPs, as revealed by an analysis of the colloids' supernatant liquids. We show that both the inner filter effect and the quenching of the dye owing to its adsorption on GNPs contribute to the observed changes in fluorescence intensity. Therefore, both factors should be properly adjusted to optimize the assay sensitivity. In particular, the low detection limit of the fluorescent DNA assay lies in the range 30-100 pM, which is close to the data reported previously for colorimetric and dynamic light scattering DNA assays.