Single-molecule fluorescence techniques on model membranes. (a) Single-vesicle content-mixing assay employing single-molecule fluorescence resonance energy transfer (smFRET) and vesicles (adapted from (60). Two distinct vesicles were reconstituted with different parts of SNARE proteins: one encapsulating dual-labeled DNA probes and the other containing complementary DNA strands. (b) smFRET with nanodiscs (adapted from (61). Each SNARE protein is fluorescently labeled and incorporated into separate nanodiscs. One nanodisc is immobilized on a polyethylene glycol (PEG)-modified glass surface, while the other is used to facilitate interaction. (c) Single-particle tracking on a free-standing lipid bilayer with customized chamber and tilted illumination (adapted from (69). The open-top chamber design allows the objective lens to be positioned close to the membrane, enhancing fluorescence detection efficiency.
