Fig. 1. The concept of STED: (a)The concept of stimulated emission; (b)the overlapping of beams.

Fig. 2. The setup of STED system.

Fig. 3. STED system based on supercontinuum laser source^[4].

Fig. 4. Simplified schematic of a STED system with a stimulated Raman-scattering light source^[5]: (a) Multicolor STED system with stimulated Raman-scattering laser source; (b) output spectrum from the SRS fiber.

Fig. 5. The intensity distributions of depletion beam^[6].

Fig. 6. Imaging curves of Confocal、traditional STED and Bessel-STED^[7]: (a) Resolution of 40 nm fluorescent beads at different depths of solid agarose samples; (b) resolution of 40 nm fluorescent beads at different depths of like-gray matter in brain tissue.

Fig. 7. The zero-order Bessel beam for excitation and the higher-order Bessel beam for depletion^[8]: (a) The intensity distribution of zero-order Bessel excitation beam in the XY plane.; (b) the intensity curve of zero-order Bessel excitation beam; (c) the intensity distribution of first-order Bessel depletion beam in the XY plane; (d) the intensity curve of first-order (black) and second-order (red)Bessel depletion beam.

Fig. 8. Alignment results of depletion and excitation beams^[9]: (a) Merged result of the excitation focus (green) and a poorly aligned depletion focus(red) using gold nanoparticles; (b) merged result of corresponding confocal (green) and STED(red) images of fluorescent beads imaged with the focus shown in (a); (c) line profiles across the dotted line in (b): confocal (green) and STED (red); (d) merged image of the excitation focus (green) and a well aligned depletion focus (red); (e) merged result of corresponding confocal (green) and STED (red) images of fluorescent beads imaged with the focus shown in (d); (f) line profiles across the solid and dashed lines in (e): confocal (green) and STED (red).

Fig. 9. Schematic of birefringent beam shaping^[10]: (a) Schematic of an easy-STED microscope using a birefringent beam shaping device; (b) merged image of the excitation beam and a depletion beam.

Fig. 10. Schematic diagram of easySLM-STED system^[11]

Fig. 11. Schematic diagram of parallel scanning STED^[17,18]: (a) Results of extended STED beam pairs; (b) optical path of the multi-point scanning STED.

Fig. 12. Schematic diagram of two-photon STED system^[24].

Fig. 13. Schematic of two-photon STED and imaging results^[25]: (a) Schematic of SW2PE-STED system; (b) 2PE and 2PE-STED imaging.

Fig. 14. Schematic diagram of two laser source dual-color STED system^[28]: (a) Dual-color STED system with two laser sources; (b) emission spectra of GFP and YFP.

Fig. 15. Schematic diagram of 3D-STED system with non-coherent wavefront modulation^[30]. Inset: combinations of phase plates and resulting STED PSFs used to achieve either ultimate lateral resolution (HR) or 3D superresolution.

Fig. 16. Schematic diagram of 4π3D-STED system^[32].

Fig. 17. Structured illumination system^[35,36]: (a) Simplified diagram of the structured illumination apparatus and spatial intensity distribution; (b) structured illumination 3D-STED system.

Fig. 18. Time-gated STED^[41]: (a) Schematic of fast-gated STED system; (b) comparison of spatial resolution for confocal, STED and gate-STED resolution.

Fig. 19. Off-line gated-STED^[44]: (a) Spatial resolutions of confocal, STED and off-line gated-STED; (b)the relationship between T_g, resolution and PSNR.

Fig. 20. STED-FLIM and Phasor-plot analysis^[45]: (a) Phasor color map of the STED-FLIM image; (b) Confocal, (c) STED·(10 mW), (d) time gate STED and (e) Phasor-plot images of fluorescent beads.

Fig. 21. Imaging results of fluorescent beads using segmentation and pSTED-SPLIT methods^[47]: (a) Schematic diagram of segmentation method; (b) schematic diagram of pSTED-SPLIT method; (c) from left to right: Confocal, conventional STED, segmentation STED, and pSTED-SPLIT imaging results of fluorescence beads.

Fig. 22. Schematic diagram of upconversion and imaging results^[48]: (a) Diagram of energy levels of upconversion nanoparticles; (b) imaging results of UCNPs.

Fig. 23. Illustration of FED theory^[53].

Fig. 24. Schematic principle of DE-STED^[55].

Fig. 25. Schematic of TIRF-STED-FCS system^[59].

Fig. 26. sSTED-FCS results of the lipid membrane^[61].