Complementary studies of lipid membrane dynamics using iSCAT and super-resolved fluorescence correlation spectroscopy.

Journal of Physics D: Applied Physics
Francesco ReinaChristian Eggeling

Abstract

Observation techniques with high spatial and temporal resolution, such as single-particle tracking based on interferometric scattering (iSCAT) microscopy, and fluorescence correlation spectroscopy applied on a super-resolution STED microscope (STED-FCS), have revealed new insights of the molecular organization of membranes. While delivering complementary information, there are still distinct differences between these techniques, most prominently the use of fluorescent dye tagged probes for STED-FCS and a need for larger scattering gold nanoparticle tags for iSCAT. In this work, we have used lipid analogues tagged with a hybrid fluorescent tag-gold nanoparticle construct, to directly compare the results from STED-FCS and iSCAT measurements of phospholipid diffusion on a homogeneous supported lipid bilayer (SLB). These comparative measurements showed that while the mode of diffusion remained free, at least at the spatial (>40 nm) and temporal (50  ⩽  t  ⩽  100 ms) scales probed, the diffussion coefficient was reduced by 20- to 60-fold when tagging with 20 and 40 nm large gold particles as compared to when using dye tagged lipid analogues. These FCS measurements of hybrid fluorescent tag-gold nanoparticle labeled lipids also revea...Continue Reading

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Citations

Aug 28, 2020·Nature Communications·Jia Hui LiHelge Ewers
Mar 8, 2019·Nature Protocols·Erdinc SezginChristian Eggeling
Nov 24, 2020·Frontiers in Cell and Developmental Biology·Richard W TaylorVahid Sandoghdar
Feb 16, 2019·The Journal of Physical Chemistry Letters·Andrey A GurtovenkoIlpo Vattulainen

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Methods Mentioned

BETA
fluorescence correlation spectroscopy
FCS
electrophoresis

Software Mentioned

FCS
MatLab
SPT
iSCAT
TrackNTrace
FoCuS
STED

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