We propose to evaluate and develop the use of metallic particles to modify fluorescence as used in DNA analysis. Fluorescence measurements are usually performed in optically homogeneously media, providing little opportunity to modify fundamental spectral properties. In contrast, recent experiments from this laboratory have shown that proximity of fluorophores to conducting metallic surfaces can increase radiative decay rates, increase quantum yields, decrease lifetimes and improve photostability. We expect these effects to substantially increase the number of photons which can be emitted by a single fluorophore. We will use silver particles and coated metallic surfaces to: 1. Examine the effects of metallic surfaces on the intrinsic emission of DNA, bases and nucleotides. The metallic particles will include silver island films, annealed films, colloids, clusters of colloids and coated metallic surfaces. 2. Examine the effects of these metallic particles on extrinsic probes commonly used to label DNA, and to explore the use of low quantum yield probes which become fluorescent near metallic surfaces. 3. Determine the effects of metallic surfaces on the rates and maximum distances for fluorescence resonance energy transfer (RET). We expect RET near metallic surfaces to extend to hundreds of A, as compared with the usual upper limit near 60 Angstroms. 4. Evaluate the use of lanthanides and transition metal-ligand complexes on DNA arrays containing metallic surfaces. These longer lived probes are highly photostable and may become useful on DNA arrays with the increased emission rate expected near metallic particles. 5. Apply the knowledge gained from Specific Aims1-4 for use on DNA arrays designed for rapid identification of antibiotic, resistant bacteria and environmental pathogens such as C. dipthariae and V. cholerae. Most experiments will be performed using metallic silver, but gold will be examined with longer wavelength DNA probes. Both intensity and lifetime measurements will be used to separate increases in the radiative rates from other effects of the sample on the fluorophore. Both one-photon and multi-photon excitation will be used. We expect this project to determine how metallic particle effects on fluorescence can be used for new approaches to DNA analysis.
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| Badugu, Ramachandram; Descrovi, Emiliano; Lakowicz, Joseph R (2014) Radiative decay engineering 7: Tamm state-coupled emission using a hybrid plasmonic-photonic structure. Anal Biochem 445:1-13 |
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