Biotechnology & Biophysics, Julius-Maximilians-University, Am Hubland, 97074, Wuerzburg, Germany.
Photochem Photobiol Sci. 2011 Apr;10(4):499-506. doi: 10.1039/c0pp00317d. Epub 2010 Dec 10.
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.
有机染料的自由基离子是高度反应性的物种，已经通过瞬态吸收光谱和脉冲辐射实验在缺氧溶液中研究了几十年。在这里，我们通过连续波 EPR、吸收和荧光实验表明，在水溶液中，巯基可以将罗丹明染料的三重态光还原为稳定的自由基阴离子，其寿命长达数小时。我们的数据表明，三重态的还原和氧对反应中间体的光诱导氧化是染料在含硫醇的水溶液中可逆光开关的一般机制，突出了分子氧在超分辨率荧光成像中的关键作用。由于细胞中含有毫摩尔浓度的巯基谷胱甘肽，并且作为副产物形成活性氧，因此我们的发现对活细胞荧光显微镜具有重要意义。
