Introduction to Turku BioImaging key technologies (see the Turku BioImaging Black Book for more information).

Superresolution Light Microscopy

Stimulated Emission Depletion (STED) microscopy is a confocal-like microscopy technique, in which the traditional diffraction resolution limit of optical microscopy can be effectively surpassed. With a STED instrument, it is possible to acquire purely optical super-resolution images, with details visible at the level of tens of nanometers.

The basic principle of STED microscopy is the use of two lasers working in a synchronized manner, in a setting that is otherwise rather similar to that of scanning confocal microscopy. One of the lasers is employed for normal fluorescence excitation, whereas the other forms a red-shifted donut-shaped beam (of zero intensity in the center), which is overlaid on the diffraction-limited fluorescence spot, and then used to deplete all fluorescence at that spot, except for the immediate center. This is achieved by exploiting an optical process termed stimulated emission depletion. By scanning the sample using a series of sub-diffraction spots, a purely optical super-resolution image can be constructed.

An STED instrument can be used in a manner very similar to that employed when other fluorescence far-field microscopes are used. Compared with, for example, electron microscopy, STED has the obvious benefit of being able to non-invasively image (living) specimens in three dimensions. Also, highly specific fluorescence labeling can be of great assistance, especially in the comparison of different biological samples. STED is the instrument of choice when the resolution of normal confocal microscopy is insufficient. Turku BioImaging offers a Leica TCS STED microscope for superresolution light microscopy.