title: Exocytotic vesicle behaviour assessed by total internal reflection fluorescence microscopy.
abstract: The regulated trafficking or exocytosis of cargo-containing vesicles to the cell surface is fundamental to all cells. By coupling the technology of fluorescently tagged fusion proteins with total internal reflection fluorescence microscopy (TIRFM), it is possible to achieve the high spatio-temporal resolution required to study the dynamics of sub-plasma membrane vesicle trafficking and exocytosis. TIRFM has been used in a number of cell types to visualize and dissect the various steps of exocytosis revealing how molecules identified via genetic and/or biochemical approaches are involved in the regulation of this process. Here, we summarize the contribution of TIRFM to our understanding of the mechanism of exocytosis and discuss the novel methods of analysis that are required to exploit the large volumes of data that can be produced using this technique.
authors.affiliation: The Garvan Institute of Medical Research, 384 Victoria Street, Sydney, New South Wales 2010, Australia.
authors.forename: James G
authors.initials: JG
authors.lastname: Burchfield
authors.forename: Jamie A
authors.initials: JA
authors.lastname: Lopez
authors.forename: Katarina
authors.initials: K
authors.lastname: Mele
authors.forename: Pascal
authors.initials: P
authors.lastname: Vallotton
authors.forename: William E
authors.initials: WE
authors.lastname: Hughes
mesh_terms: D041721:3T3-L1 Cells; D000818:Animals; D003599:Cytoskeleton / Q000378:metabolism; D005089:Exocytosis / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D010743:Phospholipids / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics; D050600:SNARE Proteins / Q000502:physiology; D022142:Secretory Vesicles / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D050857:Synaptotagmins / Q000502:physiology; D020691:rab GTP-Binding Proteins / Q000502:physiology
chemical_list: D010743:Phospholipids; D011993:Recombinant Fusion Proteins; D050600:SNARE Proteins; C485184:enhanced green fluorescent protein; D050857:Synaptotagmins; D049452:Green Fluorescent Proteins; D020691:rab GTP-Binding Proteins