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EnglishThe observation of molecular diffusion at different spatial scales, and in particular below the optical diffraction limit (<200 nm), can reveal details of the subcellular topology and its functional organization. Stimulated-emission depletion microscopy (STED) has been previously combined with fluorescence correlation spectroscopy (FCS) to investigate nanoscale diffusion (STED-FCS). However, stimulated-emission depletion fluorescence correlation spectroscopy has only been used successfully to reveal functional organization in two-dimensional space, such as the plasma membrane, while, an efficient implementation for measurements in three-dimensional space, such as the cellular interior, is still lacking. Here we integrate the STED-FCS method with two analytical approaches, the recent separation of photons by lifetime tuning and the fluorescence lifetime correlation spectroscopy, to simultaneously probe diffusion in three dimensions at different sub-diffraction scales. We demonstrate that this method efficiently provides measurement of the diffusion of EGFP at spatial scales tunable from the diffraction size down to â¼80 nm in the cytoplasm of living cells.
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| Titolo: | Measurement of nanoscale three-dimensional diffusion in the interior of living cells by STED-FCS |
| Autori: | |
| Data di pubblicazione: | 2017 |
| Rivista: | |
| Handle: | http://hdl.handle.net/11567/894906 |
| Appare nelle tipologie: | 01.01 - Articolo su rivista |
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