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119 1053-0509/03/0300-0119/0 © 2003 Plenum Publishing Corporation Five-hundred-foldenhanced fluorescence emission from fluorophore-protein coated silver fractal-like structures has been observed. These results further serve to complement our recent work on the effects of nobel metal particles with fluorophores, a relatively new phenomenon we have termed both metal-enhanced fluorescence [1] and radiative decay engineering [2,3]. We believe that this new silver-surface preparation, which results in ultrabright and photostable fluorophores, offers a new generic technology platform for increased fluorescence signal levels, with widespread potential applications to the analytical sciences, imaging, and medical diagnostics. Silver fractal-like structures have been grown on both silver electrodes and on glass slides. These structures were coated with a monolayer of human serum albumin. (HSA) protein that had been labeled with two fluorescent probes, indocyanine green (ICG) and fluorescein (FITC), respectively. Remarkable increases in fluorescence intensities were observed, typically 500-fold on the glass–silver fractal substrate and some 100-fold on the roughened silver electrodes, as compared with a control with the same substrates but with no fractal structures. In addition, dramatically decreased lifetimes and significantly increased photostabilities of the fluorescent labels were observed. Our results show that we can detect a 160-fold increase in the number of detectable photons from the labeled protein, prior to photobleaching. These exciting results are explained by the metallic surfaces modifying the radiative decay rate, G, of the fluorescent labels. Roughened silver electrodes are widely used for surface-enhanced Raman scattering (SERS) [4,5]. Although the large increases in Raman signals observed with roughened silver electrodes are thought to be due to surface-contact interactions, we questioned their use in metal-enhanced fluorescence, given that metal-enhanced fluorescence is thought to be a through-space phenomenon. We therefore reasoned that a monolayer protein spacer ( <4 nm) would provide for the through-space distance requirement of metal-enhanced fluorescence [1–3]. A constant current was passed between two silver electrodes separated by 10 mm in pure water (Fig. 1). We typically observed the growth of the fractal-like structures on the silver cathode electrode (Fig. 1, bottom). We incubated the electrode in 30 mM ICG, 60 mM HSA for 24 hr, then rinsied it with water to remove the unbound material. As a control sample, an unused electrode was also coated with ICG–HSA. Figure 2 shows the fluorescence intensity of ICG–HSA-coated roughened electrodes. The remarkable increase in fluorescence intensity, <100-fold, was also accompanied by a significantly reduced ICG lifetime, reduced .100-fold. An increase in fluorescence intensity coupled with a reduction in lifetime can only be explained by an increase in the radiative decay rate:

Fractal Silver Structures for Metal-Enhanced Fluorescence: Applications for Ultra-Bright Surface Assays and Lab-on-a-Chip-Based Nanotechnologies.