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Live mitochondria seen in unprecedented detail: photobleaching in STED microscopy overcome

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Geekz Snow
Live mitochondria seen in unprecedented detail: photobleaching in STED microscopy overcome

Moreover, by marking the biomolecules of the structure we are interested in with a specially designed fluorescent molecule, we can distinguish it from the surroundings: this is fluorescence microscopy.

Until the mid-1990s fluorescence microscopy was hampered by basic physics: due to the diffraction limit, any features on the sample closer together than about 250 nanometres would be blurred together.

But in around 1994, in a wonderful lesson teaching us that we must take care when applying fundamental physical principles, Stefan Hell discovered Stimulated Emission Depletion (STED) microscopy, which is now one of several optical microscopy approaches that achieve "super-resolution", resolution beyond the diffraction limit.

If you were to move the laser 200 nanometres in any direction, to where, in this example, no fluorescent molecules are present, the signal will certainly go dark.

So, this rather dim pixel tells us that something is present inside this sample area 200 nanometres in diameter.

First, he thought of the idea of stopping light being sent to the detector from as large an area as possible within an illuminated spot whose size matches the diffraction limit.

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