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CryoElectron Microscopy Facility Description (Word Doc file)

The CryoElectron Microscopy Facility at UCSD is housed in the first level of Bonner Hall on the Revelle College campus.  The suite occupies approximately 1700 sq. ft. and there is an additional 190 sq. ft. of wet lab space (contains a fume hood, refrigerator, light microscopes, sonicators, balance, etc) shared with another faculty member in the Division of Biology.  The cryoEM suite in Bonner Hall has nine rooms, including:

  • Three microscope rooms designed to be optimal for cryoEM experiments.  Two of these rooms are designed to contain cryo-microscopes equipped with field-emission guns.  The floors, walls and ceiling surfaces are moisture sealed so that strict temperature and humidity requirements can be maintained.  Air supply into the room is from fabric DuctSox™, air-dispersion tubes to reduce air currents in the room.  Sound dampening curtains surround the microscopes.
  • A darkroom for developing photographic film plates with a nitrogen-burst developing system
  • An equipment room that houses the microscope electronics and water chillers
  • An office for the facility manager
  • A computer room to be used for initial processing of recorded image data
  • A room that houses the dedicated Kathobar HVAC unit
  • A walk-in cooler and ante-room.  The cooler is used to prepare vitrified biological specimens.  Such conditions are required to prevent contamination of the cryogens with water vapor.  The ante-room houses some of the ancillary microscope equipment.

The facility currently houses two FEI/Philips transmission cryo-electron microscopes, including a 200kV Tecnai G2 Sphera, equipped with a LaB6 electron gun, and a 300KV Tecnai G2 Polara, equipped with a field emission gun.  The Polara has a modern stage design that allows examination of vitrified samples at either liquid helium or liquid nitrogen temperatures.  The Sphera uses standard cryo-transfer stages developed by Gatan, Inc., two of which are available for use by microscopists.  Both microscopes are completely computer controlled with the Tecnai GUI interface that runs under the Microsoft Windows 2000 operating system.  Both microscopes are also equipped with tomography image acquisition and tomographic reconstruction software.  In addition, the Leginon software package, developed by colleagues at the National Resource for Automated Molecular Microscopy in the Scripps Research Institute, is installed on both microscopes.

Features of the FEI Tecnai G2 Sphera:

  • Operates at voltages up to 200kV.
  • Cut-film holder for up to 56 films
  • Room-temperature, 70 degree tilt, sample holder
  • Gatan Ultrascan 1000 UHS CCD camera designed for a 200kV electron source.  The camera has a 4 megapixel (2k X 2k), Peltier-cooled, CCD chip and is equipped with an ultra-high sensitivity phosphor scintillator.  The camera is mounted on-axis under the microscope column and is connected via an optical cable to the microscope computer.
  •  Gatan 676 on-axis, intensified TV camera with a YAG scintillator and a 12 in, monochrome monitor.
  • Cryo package: includes an improved anticontaminator, Low-Dose software, and cryo-cycle protection for the ion getter pump
  • Gatan Digital Micrograph software: operates the CCD camera and permits various levels of analysis of recorded images
  • Two, NEC 20” LCD color monitors
  • Gatan HREM Autotune software: automatically sets image defocus and stigmates CCD images
  • FEI Xplore3D tomographic software
  • SerialEM tomographic software (D. Mastronarde, University of Colorado at Boulder)
  • Tecnai Scripting and Photomontage software
  • NRAMM Leginon automated data acquisition software
  • SerialEM tomographic software (D. Mastronarde, University of Colorado at Boulder)
  • Two Gatan, 70-degree tilt cryo-transfer holders and one cryo-transfer station
  • Gatan Dry Pumping Station: used to pump out cryo-holders for improved performance
  • Miscellaneous ancillary equipment: e.g. a Haskris water circulator; ZEM water circulator; film desiccator; etc.

Features of the FEI Tecnai G2 Polara

  • Electron gun is a Schottky field emitter. 
  • Operates at voltages up to 300kV.
  • Multi-specimen transfer device: has 6 storage positions for cartridges that hold 3 mm EM grids
  • Cryoworkstation: for mounting EM grids into the cartridges and to load the cartridges into the multi-specimen transfer device.  The workstation has a loading area, a vacuum system and an airlock for docking to the transfer device.  The workstation can be rolled out of the way when not needed.
  • Improved vacuum system: consists of a rotary pump and buffer tank, an oil diffusion pump, a turbo-molecular pump, and four ion getter pumps
  • Computer-controlled, ultra-stable goniometer that can be cooled down to either liquid nitrogen or helium temperatures with the ability to tilt up to 70 degrees.
  • Cut-film holder for up to 56 films
  • Gatan Ultrascan 4000 UHS CCD camera designed for a 300kV electron source.  The camera has a 16 megapixel (4k X 4k), Peltier-cooled, CCD chip and is equipped with an ultra-high sensitivity phosphor scintillator.  The camera is mounted on-axis under the microscope column and is connected via an optical cable to the computer of the microscope.
  • Cryo package: includes an improved anticontaminator “cryobox”, Low-Dose software, and cryo-cycle protection for the ion getter pumps
  • Gatan Digital Micrograph software: operates the CCD camera and permits various levels of analysis of recorded images
  • Two, NEC 20” LCD color monitors
  • Gatan HREM Autotune software: automatically sets image defocus and stigmates CCD images
  • FEI Xplore3D tomographic software
  • SerialEM tomographic software (D. Mastronarde, University of Colorado at Boulder)
  • Tecnai Scripting and Photomontage software
  • NRAMM Leginon automated data acquisition software
  • SerialEM tomographic software (D. Mastronarde, University of Colorado at Boulder)
  • Miscellaneous ancillary equipment: e.g. a Haskris water circulator, Zem water circulator; film desiccator; etc.

Auxilliary Equipment: (in the Bonner facility or in the nearby Natural Sciences Building)

  • Emitech K950X carbon evaporator equipped with a K150 film thickness monitor and a K350 glow discharge device
  • Two manual, freeze-plunge devices for preparing vitrified samples
  • Folded optical diffractometer: used both for assessing quality of micrographs and as a teaching tool to help new staff learn principles of diffraction theory
  • Nikon Super Coolscan 8000ED film scanner: for digitizing micrographs at 4000 dpi pixel resolution.
  • Epson Perfection 4870 Photo flatbed scanner, 4800 X 9600dpi, 48bit color depth
  • Two Dell Latitude D610 computers each with 1 GB memory, 80 GB hard drive, 14 in flat panel displays, Linux Centos OS.
  • StorageWare SA214 2U Dual Xeon SATA server with one 80 GB SATA hard drive and six 250 GB  SATA hard drives
  • Hewlett Packard LaserJet 4350n printer, 1200 dpi
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