Institute of Physical and Theoretical Chemistry, Johann Wolfgang Goethe-University, Max-von-Laue-Str. 7, 60438 Frankfurt (Germany).
Chemphyschem. 2014 Mar 17;15(4):637-41. doi: 10.1002/cphc.201300874. Epub 2013 Dec 2.
In spite of their relatively low fluorescence quantum yield, cyanine dyes such as Cy3, Cy5, or Cy7 are widely used in single-molecule fluorescence applications due to their high extinction coefficients and excellent photon yields. We show that the fluorescence quantum yield and lifetime of red-emitting cyanine dyes can be substantially increased in heavy water (D2 O) compared with water (H2 O). We find that the magnitude of the quantum yield increase in D2 O scales with the emission wavelength, reaching a particularly high value of 2.6-fold for the most red-emitting dye investigated, Cy7. We further demonstrate a higher photon yield in single-molecule superresolution experiments in D2 O compared to H2 O, which leads to an improved localization precision and hence better spatial resolution. This finding is especially beneficial for biological applications of fluorescence microscopy, which are typically carried out in aqueous media and which greatly profit from the red spectral range due to reduced cellular auto-fluorescence.
尽管量子产率相对较低，但由于其高消光系数和优异的光子产率，菁染料（如 Cy3、Cy5 或 Cy7）在单分子荧光应用中得到了广泛应用。我们表明，与水（H2O）相比，重水（D2O）中红色发射菁染料的荧光量子产率和寿命可以大大提高。我们发现，D2O 中量子产率的增加幅度与发射波长成正比，对于研究的最红色发射染料 Cy7，达到了特别高的 2.6 倍。我们进一步证明，在单分子超分辨率实验中，D2O 中的光子产率高于 H2O，这导致定位精度提高，从而获得更好的空间分辨率。这一发现对于荧光显微镜的生物学应用尤其有益，因为荧光显微镜的生物学应用通常在水相介质中进行，并且由于细胞自发荧光减少，红色光谱范围非常有利。
