Fig. 4: Investigation of endosomal vesicle interaction with etSTED.

This shows an etSTED experiment in hippocampal neurons expressing CD63-EGFP and using the vesicle_proximity analysis pipeline. a, Schematic diagram of an endosomal vesicle interaction process, with the increasing proximity due to one labeled vesicle moving towards another stationary vesicle as labeled with CD63-EGFP. b, Schematic diagram of the experiment timeline of one widefield frame. c, Example of a triggering widefield frame. d, Two representative events shown in widefield zooms with three frames leading up to the event (right frame). The green box indicates the area of the STED scan. e, Triggered 2.8 Hz STED timelapses of endosomal vesicle dynamics where sphingosyl PE (sphingosyl-PE_Abberior STAR RED) is labeled. n = 123 events, n = 23 cells. Scale bars, 10 µm (c), 2 µm (d), 250 nm (e). The vesicles involved in the event are marked by symbols (asterisks and arrowhead).