Executive Summary : | Despite many applications, the fluorescence methods including Förster resonance energy transfer (FRET) still face the challenge of low signal to noise ratio and quick photobleaching of the fluorophores which limits the fluorescence methods to some handful of fluorophores with high fluorescence quantum yield. Furthermore, FRET detection is limited to 10 nm distance range only, which prevents its use to probe the structural dynamics of large proteins. The proposal aims to overcome these challenges of fluorescence methods using plasmonic nanoantenna. In particular, the proposal would like to use the intense localized electromagnetic hotspot offered by the nanoantenna to boost the fluorescence counts and long distance FRET of the fluorescent probes. For this purpose, the proposal aims to use the anisotropic dimer nanogap antenna where two anisotropic metal nanoparticles (NPs) will be separated by a few nm gap. This type of nanoantenna will generate a highly intense hotspot in the gap due to coupling of the localized surface plasmon resonance (LSPR) of individual metal NPs. Anisotropic dimer nanogap antenna will be fabricated through cheaper colloidal chemical synthesis which produces monocrystalline nanostructures with superior plasmonic effect than the lithographically fabricated nanostructures. However, it still lacks control and reproducibility in precisely assembling the metal NPs to form an effective optical antenna. The proposal will utilize DNA based self assembly to overcome this challenge. In this approach, first individual anisotropic Au and Ag NPs building blocks will be prepared using the seed mediated growth method. Next, these building blocks will be conjugated to a thiol functionalized DNA single strands. Finally, the dimer nanogap antenna will be fabricated through the hybridization of the complimentary strands of DNA. The gap size of the nanoantenna will be varied by taking DNA of different lengths. To place the fluorophore exactly at the nanoantenna hotspot one of the DNA single strands in the DNA will be labelled with the fluorophore. A range of fluorophores with emission maxima varying from visible to near IR will be used. Au nanoantenna will be used to boost the fluorescence of the red to near IR emitting fluorophores while Ag nanoantenna will be used to increase the fluorescence of blue to yellow emitting fluorophores due to their superior plasmonic properties in these spectrum. The fluorescence counts and lifetime will be monitored as a function of nanoantenna gap size to find the optimum gap size for fluorescence enhancement. To boost the long range FRET, the DNA will be labelled with donor (D) and acceptor (A) fluorophore at suitable positions to have the D-A separation distance greater than 10 nm. FRET enhancement will be monitored as a function of D-A separation distance. Overall, the project will establish general guidelines to obtain optimum enhancement of fluorescence using nanoplasmonics. |