A Brief Introduction to Super-resolution Microscopy

'Super-resolution microscopy' or 'imaging' generally refers to any microscopic technique capable of performing images with resolution below the diffraction limit.

By Dr Jakob Bierwagen, AHF analyentechnik AG

All superresolution methods use techniques to distinguish fluorophores by different states. These can be, for example, the S0 and S1 states or cis and trans or differentiation by different bright and dark states. The well-known methods for superresolution are: STED (stimulated emission depletion) and the other RESOLFT methods, STORM, PALM, GSDIM and SOFI, as well as SIM, SSIM and MINFLUX. One can divide these methods into the following categories:

STED and the other RESOLFT- methods are based on purely physical methods that increase the resolution, e.g., stimulated emission in STED.

STORM, PALM, GSDIM, SOFI and their derivatives are based on (computational) localization of single fluorophores, which results in an superresolved image by superposition of many images.

SIM and SSIM mainly use deconvolution methods to calculate the "true" object from the image. For this purpose, the sample is illuminated in a structured manner.

Finally, MINFLUX uses RESOLFT techniques for localisation as well as techniques from the localization methods for switching and thus achieves molecular resolution (2-5 nm) even in biological samples closing the gap to electron microscopy.