
pmid: 29454082
Combining stimulated emission depletion and fluorescence correlation spectroscopy (STED-FCS) provides a powerful and sensitive tool for studying the molecular dynamics in live cells with high spatio-temporal resolution. STED-FCS gives access to molecular diffusion characteristic at the nanoscale occurring within short period of times. However due to the incomplete suppression of fluorescence in the STED process, the STED-FCS point spread function (PSF) deviates from a Gaussian shape and challenges the analysis of the auto-correlation curves obtained by FCS. Here, we model the effect of the incomplete fluorescence suppression in STED-FCS experiments and propose a new fitting model improving the accuracy of the diffusion times and average molecule numbers measurements. The implementation of a STED module with pulsed laser source on a commercial confocal/FCS microscope allowed us to apply the STED-background corrected model to fit the STED-FCS measurements. The experimental results are in good accordance with the theoretical analysis both for the number of molecules and the diffusion time which decrease accordingly with the STED power.
[SDV.IMM] Life Sciences [q-bio]/Immunology, Intravital Microscopy, Lasers, Fluorescence, Laser Scanning Cytometry, Diffusion, Actin Cytoskeleton, Spectrometry, Fluorescence, Microscopy, Fluorescence, Models, Chemical, COS Cells, Chlorocebus aethiops, Image Processing, Computer-Assisted, Animals, Software
[SDV.IMM] Life Sciences [q-bio]/Immunology, Intravital Microscopy, Lasers, Fluorescence, Laser Scanning Cytometry, Diffusion, Actin Cytoskeleton, Spectrometry, Fluorescence, Microscopy, Fluorescence, Models, Chemical, COS Cells, Chlorocebus aethiops, Image Processing, Computer-Assisted, Animals, Software
| selected citations These citations are derived from selected sources. This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically). | 8 | |
| popularity This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network. | Average | |
| influence This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically). | Average | |
| impulse This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network. | Top 10% |
