We studied the dependence of single-molecule fluorescence on the size of nearby metal particles. The silver particles were synthesized with average diameters of metal cores being 5, 20, 50, 70, and 100 nm, respectively. A single-stranded oligonucleotide was chemically bound to a single silver particle and a Cy5-labeled complementary single-stranded oligonucleotide was hybridized with the particle-bound oligonucleotide. The space between the fluorophore and metal core was separated by a rigid hybridized DNA duplex of 8 nm length. The single fluorescence images and intensity traces were recorded by scanning confocal microscopy. The single fluorophore-labeled 50 nm silver particles displayed the most enhanced intensity, a factor of 17-fold increase relative to the free fluorophores in the absence of metal. Numerical simulations by the finite-difference time-domain (FDTD) method and results from Mie theory were used to compare with the experimental results. The 50 nm silver particles were also labeled by multiple fluorophores. The fluorescence intensity of multiple fluorophore-labeled metal particles increases dramatically with the loading number and reached 400-fold relative to the free single fluorophore when the loading number of fluorophore per metal particle was 50. The fluorophore also displayed better photostability when binding on the metal particle. These results can aid us to develop novel nanoscale fluorophores for clinical diagnostics and bioassay.
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