Single-stranded oligonucleotides stabilize highly fluorescent Ag nanoclusters, with emission colors tunable via DNA sequence. We utilized DNA microarrays to optimize these scaffold sequences for creating nearly spectrally pure Ag nanocluster fluorophores that are highly photostable and exhibit great buffer stability. Five different nanocluster emitters have been created with tunable emission from the blue to the near-IR and excellent photophysical properties. Ensemble and single molecule fluorescence studies show that oligonucleotide encapsulated Ag nanoclusters exhibit significantly greater photostability and higher emission rates than commonly used cyanine dyes.

This Review summarizes the latest advances in the field of rylene dyes and rylene nanoemitters for applications in photonics, and describes the influence of the dye design on the optical properties, the self-assembly, the molecular interactions, as well as the labeling specificity of the compounds. The interplay between tailored (macro)molecular design and bulk/single-molecule spectroscopy enables complex processes to be explained, for example, the kinetics of energy-transfer processes or (bio)catalysis. Such investigations are essential for the ultimate design of optimized nanoemitters, and require a close cooperation between spectroscopists and preparative organic chemists.

The water-soluble, near-IR-emitting DNA-encapsulated silver nanocluster presented herein exhibits extremely bright and photostable emission on the single-molecule and bulk levels. The photophysics have been elucidated by intensity-dependent correlation analysis and suggest a heavy atom effect of silver that rapidly depopulates an excited dark level before quenching by oxygen, thereby conferring great photostability, very high singlemolecule emission rates, and essentially no blinking on experimentally relevant time scales (0.1 to >1,000 ms). Strong antibunching is observed from these biocompatible species, which emit >10 9 photons before photobleaching. The significant dark-state quantum yield even enables bunching from the emissive state to be observed as a dip in the autocorrelation curve with only a single detector as the dark state precludes emission from the emissive level. These species represent significant improvements over existing dyes, and the nonpower law blinking kinetics suggest that these very small species may be alternatives to much larger and strongly intermittent semiconductor quantum dots.correlation ͉ photophysics ͉ silver nanoclusters ͉ single-molecule spectroscopy ͉ fluorescence intermittency W hile myriad dyes exist with varying photophysical properties (1, 2), organic dye-based single-molecule and even bulk in vivo imaging dynamics studies suffer from low probe brightness, poor photostability (3), and oxygen sensitivity (4). Advances in nanotechnology such as the use of quantum dots (5, 6) have ameliorated some of these issues but at the cost of toxicity (7), broad excitation (8, 9), power-law blinking (10-12), and large probe size (13,14). While quantum dots are readily excited with low-intensity sources, their fluorescence exhibits intermittency on all time scales (10-12), thereby causing problems when used for tracking or imaging studies. Arising from Auger processes (15), these photophysical dynamics are apparent at all excitation intensities and appear without characteristic times. While functionalization, large size (Ϸ10-20 nm in diameter), and cellular uptake are potential problems, the strong nonmolecular power-law fluorescence intermittency is a major drawback of these materials as single-molecule reporters (10-12). Recently, Ϸ35-nm-sized fluorescent nanodiamonds have also been reported as single-molecule emitters, but these also raise concerns about label size (16). Consequently, for both in vitro and in vivo single-molecule studies, fluorophores with high emission rates and excellent photostability must be identified that are completely devoid of blinking on all relevant time scales, while maintaining small overall sizes.By combining the virtues of chemistry and nanotechnology, we have developed few-atom, molecular-scale noble metal nanoclusters as a class of emitters that simultaneously exhibit bright, highly polarizable discrete transitions, good photostability, and small size, all within biocompatible scaffolds (17-20). Recent observations that DNA encapsulates Ag nanoclust...

Individual multichromophoric dendrimer molecules, bearing eight perylenecarboximide chromophores at the rim, immobilized in a thin polyvinylbutyral (PVB) film were studied by far-field fluorescence microscopy. Fluorescence intensity trajectories as a function of time (transients), spectra, and decay traces were recorded separately or simultaneously. For comparison, similar measurements have been performed on a model compound containing one perylenecarboximide chromophore. Collective on/off jumps of the fluorescence intensity were observed for single dendrimer molecules, resembling previously reported collective jumps for the emission of single light-harvesting antenna systems. Spectra and decays of both non-interacting and dimer-like interacting chromophoric sites could be distinguished within an individual dendrimer. Transitions between the different spectral forms and decay times, observed for a single molecule, underline the dynamic character of the interactions among the chromophores. Evidence for a stepwise bleaching process of the multichromophoric system was found. Furthermore, the single-molecule data incontestably prove the assumptions stated in the ensemble model.