Live-cell STED microscopy with genetically encoded biosensor

Of the various super-resolution techniques, stimulated emission depletion (STED) microscopy achieves the best temporal resolution at high spatial resolution, enabling live-cell imaging beyond the diffraction limit. However, STED and most other super-resolution imaging methods utilize a particular type of information extractable from the raw data, namely the positions of fluorophores. To expand on the use of super-resolution techniques, we report here the live-cell STED microscopy of a dynamic biosensor. Using the fluorescent H₂O₂ sensor HyPer2 for subdiffraction imaging, we were able not only to image filaments with superior resolution by localizing emission but also to trace H₂O₂ produced within living cell by monitoring brightness of the probe. STED microscopy of HyPer2 demonstrates potential utility of FP-based biosensors for super-resolution experiments in situ and in vivo.