Abstract
Photophysical phenomena leading to blinking and irreversible photobleaching represent a major obstacle that limits the utility of organic fluorophores in fluorescence-based techniques involving the detection of small numbers of molecules. In recent work, we demonstrated that the coordination of paramagnetic transition metals (e.g. Mn2+) to DNA induces intersystem-crossing in dyes covalently attached to the nucleic acid. This results in fluorescence quenching, triplet blinking and accelerated photobleaching. The increase in triplet formation in the presence of manganese was demonstrated using transient absorption techniques and fluorescence correlation spectroscopy for a series of rhodamine and cyanine dyes, including TAMRA, Cy3 and Cy5. These results are particularly relevant for single-molecule or fluorescence correlation spectroscopy experiments aimed to study enzymes that act on DNA, where Mn2+ is used to relax the sequence-specificity of enzymes that catalyze phosphoryl transfer reactions (e.g. polymerases and restriction endonucleases).
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