Fig. 2a–e.

Reversible saturable optical fluorescence transition (RESOLFT) microscopy. a A fluorophore can be switched reversibly between a dark state and a bright state using light. A special illumination pattern can be applied that reduces the effective size of the point spread function (PSF), for example, by stimulated emission depletion (STED). b Sketch of the scanning procedure. The excitation beam is overlayed with the donut-shaped depletion beam with zero intensity in the center. Thus, all molecules further away from the center (red dots) are efficiently deexcited by stimulated emission before they fluoresce; only those molecules close to the center emit fluorescence (green dots). c STED image of vimentin filaments in PtK2 cells labeled with primary and secondary antibodies. d Confocal image of the region marked by the white frame in (c). e STED image of the same region, the cross-section marked by a white bar features a width of 68 nm; scale bars 1 μm. Reprinted with kind permission (Moneron et al. 2010)