title: Photoinduced formation of reversible dye radicals and their impact on super-resolution imaging.
abstract: Radical ions of organic dyes are highly reactive species and have been studied for decades by transient absorption spectroscopy and pulse radiolysis experiments in oxygen-depleted solution. Here we show by continuous wave EPR, absorption, and fluorescence experiments that the triplet state of rhodamine dyes can be photoreduced by thiols to form stable radical anions in aqueous solution with a lifetime of up to several hours. Our data demonstrate that reduction of the triplet state and photoinduced oxidation of reactive intermediates by oxygen represents a general mechanism for reversible photoswitching of dyes in aqueous thiol-containing solutions highlighting the key role of molecular oxygen for super-resolution fluorescence imaging. Since cells contain the thiol glutathione at millimolar concentrations and reactive oxygen species are formed as side products our findings are of consequence for live cell fluorescence microscopy.
authors.affiliation: Biotechnology & Biophysics, Julius-Maximilians-University, Am Hubland, 97074, Wuerzburg, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: van de Linde
authors.forename: Ivan
authors.initials: I
authors.lastname: Krstić
authors.forename: Thomas
authors.initials: T
authors.lastname: Prisner
authors.forename: Sören
authors.initials: S
authors.lastname: Doose
authors.forename: Mike
authors.initials: M
authors.lastname: Heilemann
authors.forename: Markus
authors.initials: M
authors.lastname: Sauer
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004578:Electron Spin Resonance Spectroscopy; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000528:radiation effects; D005609:Free Radicals / Q000737:chemistry / Q000378:metabolism*; D005978:Glutathione / Q000737:chemistry; D008027:Light; D008280:Magnetics; D008856:Microscopy, Fluorescence; D010084:Oxidation-Reduction; D012235:Rhodamines / Q000737:chemistry* / Q000528:radiation effects; D013438:Sulfhydryl Compounds / Q000737:chemistry
chemical_list: D005456:Fluorescent Dyes; D005609:Free Radicals; D012235:Rhodamines; D013438:Sulfhydryl Compounds; D005978:Glutathione