Abstract
Photobleaching and blinking of fluorophores pose fundamental limitations on the information content of single-molecule fluorescence measurements. Photoinduced blinking of Cy5 has hampered many previous investigations using this popular fluorophore. Here we show that Trolox in combination with the enzymatic oxygen-scavenging system eliminates Cy5 blinking, dramatically reduces photobleaching and improves the signal linearity at high excitation rates, significantly extending the applicability of single-molecule fluorescence techniques.
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Acknowledgements
We thank J. Liu for introducing Trolox to us and the US National Institutes of Health for grants.
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Supplementary Fig. 1
Single molecule FRET experiments in the presence of 2 mM Trolox or 143 mM BME for the construct of Figure 2d. (PDF 38 kb)
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Rasnik, I., McKinney, S. & Ha, T. Nonblinking and long-lasting single-molecule fluorescence imaging. Nat Methods 3, 891–893 (2006). https://doi.org/10.1038/nmeth934
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DOI: https://doi.org/10.1038/nmeth934
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