title: ADVANCED IMAGING. Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics.
abstract: Super-resolution fluorescence microscopy is distinct among nanoscale imaging tools in its ability to image protein dynamics in living cells. Structured illumination microscopy (SIM) stands out in this regard because of its high speed and low illumination intensities, but typically offers only a twofold resolution gain. We extended the resolution of live-cell SIM through two approaches: ultrahigh numerical aperture SIM at 84-nanometer lateral resolution for more than 100 multicolor frames, and nonlinear SIM with patterned activation at 45- to 62-nanometer resolution for approximately 20 to 40 frames. We applied these approaches to image dynamics near the plasma membrane of spatially resolved assemblies of clathrin and caveolin, Rab5a in early endosomes, and α-actinin, often in relationship to cortical actin. In addition, we examined mitochondria, actin, and the Golgi apparatus dynamics in three dimensions.
authors: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA 20147, USA.
authors: Dong
authors: D
authors: Li
authors: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA 20147, USA.
authors: Lin
authors: L
authors: Shao
authors: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA 20147, USA.
authors: Bi-Chang
authors: BC
authors: Chen
authors: Key Laboratory of RNA Biology and Beijing Key Laboratory of Noncoding RNA, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China. College of Life Sciences, Central China Normal University, Wuhan 430079, Hubei, China.
authors: Xi
authors: X
authors: Zhang
authors: Key Laboratory of RNA Biology and Beijing Key Laboratory of Noncoding RNA, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors: Mingshu
authors: M
authors: Zhang
authors: Coleman Technologies, 5131 West Chester Pike, Newtown Square, PA 19073, USA.
authors: Brian
authors: B
authors: Moses
authors: Coleman Technologies, 5131 West Chester Pike, Newtown Square, PA 19073, USA.
authors: Daniel E
authors: DE
authors: Milkie
authors: Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors: Jordan R
authors: JR
authors: Beach
authors: Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors: John A
authors: JA
authors: Hammer
authors: Department of Cell Biology and Pediatrics, Harvard Medical School and Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA.
authors: Mithun
authors: M
authors: Pasham
authors: Department of Cell Biology and Pediatrics, Harvard Medical School and Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA.
authors: Tomas
authors: T
authors: Kirchhausen
authors: Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA. National High Magnetic Field Laboratory and Department of Biological Science, Florida State University, Tallahassee, FL 32310, USA.
authors: Michelle A
authors: MA
authors: Baird
authors: National High Magnetic Field Laboratory and Department of Biological Science, Florida State University, Tallahassee, FL 32310, USA.
authors: Michael W
authors: MW
authors: Davidson
authors: Key Laboratory of RNA Biology and Beijing Key Laboratory of Noncoding RNA, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors: Pingyong
authors: P
authors: Xu
authors: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA 20147, USA. betzige@janelia.hhmi.org.
authors: Eric
authors: E
authors: Betzig
mesh_terms: D000185:Actinin / Q000032:analysis; D000199:Actins / Q000032:analysis; D000818:Animals; D002460:Cell Line; D002966:Clathrin / Q000032:analysis; D022163:Clathrin-Coated Vesicles / Q000737:chemistry / Q000648:ultrastructure; D003034:Coated Pits, Cell-Membrane / Q000737:chemistry / Q000648:ultrastructure; D003599:Cytoskeleton / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D004705:Endocytosis; D011992:Endosomes / Q000737:chemistry / Q000648:ultrastructure; D006056:Golgi Apparatus / Q000648:ultrastructure; D007091:Image Processing, Computer-Assisted; D021621:Imaging, Three-Dimensional / Q000295:instrumentation / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D008928:Mitochondria / Q000737:chemistry / Q000648:ultrastructure; D015388:Organelles / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D020696:rab5 GTP-Binding Proteins / Q000032:analysis
chemical_list: D000199:Actins; D002966:Clathrin; D000185:Actinin; D020696:rab5 GTP-Binding Proteins