title: A hybrid high-speed atomic force-optical microscope for visualizing single membrane proteins on eukaryotic cells.
abstract: High-speed atomic force microscopy is a powerful tool for studying structure and dynamics of proteins. So far, however, high-speed atomic force microscopy was restricted to well-controlled molecular systems of purified proteins. Here we integrate an optical microscopy path into high-speed atomic force microscopy, allowing bright field and fluorescence microscopy, without loss of high-speed atomic force microscopy performance. This hybrid high-speed atomic force microscopy/optical microscopy setup allows positioning of the high-speed atomic force microscopy tip with high spatial precision on an optically identified zone of interest on cells. We present movies at 960 ms per frame displaying aquaporin-0 array and single molecule dynamics in the plasma membrane of intact eye lens cells. This hybrid setup allows high-speed atomic force microscopy imaging on cells about 1,000 times faster than conventional atomic force microscopy/optical microscopy setups, and allows first time visualization of unlabelled membrane proteins on a eukaryotic cell under physiological conditions. This development advances high-speed atomic force microscopy from molecular to cell biology to analyse cellular processes at the membrane such as signalling, infection, transport and diffusion.
authors.affiliation: U1006 INSERM, Université Aix-Marseille, Parc Scientifique et Technologique de Luminy, 163 avenue de Luminy, Marseille, France.
authors.forename: Adai
authors.initials: A
authors.lastname: Colom
authors.forename: Ignacio
authors.initials: I
authors.lastname: Casuso
authors.forename: Felix
authors.initials: F
authors.lastname: Rico
authors.forename: Simon
authors.initials: S
authors.lastname: Scheuring
mesh_terms: D000818:Animals; D020346:Aquaporins / Q000737:chemistry* / Q000378:metabolism; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure*; D018914:Cells, Immobilized; D004926:Escherichia coli / Q000737:chemistry / Q000648:ultrastructure*; D005136:Eye Proteins / Q000737:chemistry* / Q000378:metabolism; D007908:Lens, Crystalline / Q000378:metabolism / Q000648:ultrastructure*; D018625:Microscopy, Atomic Force / Q000295:instrumentation / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D056004:Molecular Dynamics Simulation; D061251:Primary Cell Culture; D012756:Sheep; D034561:Sheep, Domestic; D059008:Time-Lapse Imaging
chemical_list: D020346:Aquaporins; D005136:Eye Proteins; C030328:aquaporin 0