PT  - JOURNAL ARTICLE
AU  - Aurélien Barbotin
AU  - Iztok Urbančič
AU  - Silvia Galiani
AU  - Christian Eggeling
AU  - Martin Booth
AU  - Erdinc Sezgin
TI  - z-STED imaging and spectroscopy to investigate nanoscale membrane structure and dynamics
AID  - 10.1101/2019.12.28.889923
DP  - 2019 Jan 01
TA  - bioRxiv
PG  - 2019.12.28.889923
4099  - http://biorxiv.org/content/early/2019/12/28/2019.12.28.889923.short
4100  - http://biorxiv.org/content/early/2019/12/28/2019.12.28.889923.full
AB  - Super-resolution STED microcopy provides optical resolution beyond the diffraction limit. The resolution can be increased laterally (xy/2D) or axially (z/3D). 2D STED has been extensively used to elucidate the nanoscale membrane structure and dynamics, via imaging or combined with spectroscopy techniques such as fluorescence correlation spectroscopy (FCS) and spectral imaging. On the contrary, z-STED has not been used in this context. Here, we show that a combination of z-STED with FCS or spectral imaging enables us to see previously unobservable aspects of cellular membranes. We show that thanks to an axial resolution of approximately 100 nm, z-STED can be used to distinguish axially close-by membranes, early endocytic vesicles or tubular membrane structures. Combination of z-STED with FCS and spectral imaging showed diffusion dynamics and lipid organization in these structures, respectively.