Switching of Organic Fluorophores by Glycerol‐sulfite Interactions for Single‐Molecule Super‐Resolution Microscopy**

Abstract

AbstractPhotoinduced off‐switching of organic fluorophores is used in super‐resolution microscopy to separate and localize single fluorescent molecules. However, this method typically relies on the use of complex imaging buffers that make use of enzymatic oxygen scavengers to remove oxygen and primary thiols to reversibly reduce excited fluorophores to a non‐fluorescent state. Here, we show that the interactions between sodium sulfite and glycerol can also be used for efficient switching of common organic fluorophores. The addition of glycerol to the imaging buffer allows us to tune the dye switching from intermittent fluorescence to the off‐state. We additionally show that sodium sulfite‐glycerol‐based buffers are especially useful for imaging at low laser illumination intensities in high‐refractive index solutions and samples embedded in agarose gel. This significantly simplifies the implementation of different super‐resolution microscopy modalities such as direct STochastic Optical Reconstruction Microscopy (dSTORM) and Super‐resolution Optical Fluctuation Imaging (SOFI).