title: The contribution of reactive oxygen species to the photobleaching of organic fluorophores.
abstract: Photoexcitation of fluorophores commonly used for biological imaging applications generates reactive oxygen species (ROS) which can cause bleaching of the fluorophore and damage to the biological system under investigation. In this study, we show that singlet oxygen contributes relatively little to Cy5 and ATTO 647N photobleaching at low concentrations in aqueous solution. We also show that Cy5 generates significantly less ROS when covalently linked to the protective agents, cyclooctatetraene (COT), nitrobenzyl alcohol (NBA) or Trolox. Such fluorophores exhibit enhanced photostability both in bulk solutions and in single-molecule fluorescence measurements. While the fluorophores ATTO 647N and ATTO 655 showed greater photostability than Cy5 and the protective-agent-linked Cy5 derivatives investigated here, both of ATTO 647N and ATTO 655 generated singlet oxygen and hydroxyl radicals at relatively rapid rates, suggesting that they may be substantially more phototoxic than Cy5 and its derivatives.
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University.|Tri-Institutional Training Program in Chemical Biology, 1300 York Avenue, New York, NY 10065, USA.
authors.forename: Qinsi
authors.initials: Q
authors.lastname: Zheng
authors.affiliation: Department of Chemistry, Columbia University, New York, NY 10027, USA.
authors.forename: Steffen
authors.initials: S
authors.lastname: Jockusch
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Zhou
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University.|Tri-Institutional Training Program in Chemical Biology, 1300 York Avenue, New York, NY 10065, USA.
authors.forename: Scott C
authors.initials: SC
authors.lastname: Blanchard
mesh_terms: D005456:Fluorescent Dyes / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D009930:Organic Chemicals / Q000737:chemistry*; D038761:Photobleaching; D017382:Reactive Oxygen Species / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D009930:Organic Chemicals; D017382:Reactive Oxygen Species