Mikroskopie

The resolution of an optical microscope is fundamentally limited by diffraction. In a conventional wide-field fluorescence microscope, the resolution limit is around 200nm. However, modern super-resolution methods can bypass this limit. Pointillistic imaging techniques like PALM and STORM do so by precisely localizing each individual molecules in a sample. In contrast, STED uses the stimulated emission process driven to saturation to dramatically reduce the size of the region in the sample which is capable of spontaneously emitting fluorescence. SIM illuminates the sample with a pattern, typically the image of a grating. This allows to computationally remove the out-of-focus blur, a method known as optical sectioning SIM. Furthermore, frequency mixing of the illumination pattern with the sample caused by the moiré effect results in a down-modulation of fine sample detail into the frequency-support of the detection optical transfer function. High-resolution SIM achieves typically a two-fold lateral resolution enhancement. This was further improved by utilizing a non-linear sample response to the illumination light in SIM. Recent developments of the method allow fast, multicolor, and three dimensional high-resolution live cell imaging.