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Resource for the Visualization of Biological Complexity (RVBC)

IVEM: JEOL JEM-4000FX Intermediate-Voltage Electron Microscope

Main features

  • 100-400 keV accelerating voltage
  • Post-column energy filter
  • Software for automated tomographic tilt-series collection
  • 2k CCD camera for image recording
  • 1k TV-rate CCD camera for specimen tracking
  • Analytical capability: EFTEM, STEM, EELS, EDX

Characteristics

Objective lens
Focal length -- 3.1 mm
Chromatic aberration -- 2.2 mm
Spherical aberration -- 1.8 mm
Resolution
TEM lattice -- 0.14 nm
TEM point to point -- 0.22 nm
STEM edge to edge -- 1.0 nm
SEM edge to edge -- 2.0 nm

Energy filter

Gatan GIF2002 with Gatan MSC 794 2k CCD camera: Zero-loss filtered imaging is used to improve contrast of frozen-hydrated specimens. Energy-filtered imaging is also used to acquire compositional maps of elements present in the specimen. The energy filter is integrated with the software for automated tomographic tilt-series collection. The CCD camera is normally binned to 1024x1024 pixels, yielding a CCD element size of 60 µm, which fully accommodates the point-spread function of a special thick phosphor scintillator. This configuration gives optimum performance for imaging dose-sensitive specimens at high accelerating voltage.

Specimen holders

  • Cryo-transfer
    • Gatan 626 single-tilt
    • Gatan CT3500TR tilt-rotation
  • Room temperature
    • Tilt-rotation
    • Dual-grid double-tilt with Faraday cage
    • Double-tilt, low background

Auxiliary CCD camera

A TVIPS FastScan 1k CCD camera, with a custom phosphor scintillator, is mounted off-axis next to the energy filter. This camera is used to search and track specimens at lower magnification than the main CCD camera, and at very low dose. It is also used to record electron diffraction patterns of frozen-hydrated specimens to asses the state of the ice.

Tomography software

Collection of tomographic tilt series is carried out using automation software written in-house. The software integrates the energy filter, the main CCD camera, the TVIPS CCD camera, the specimen stage, and the EM itself. The tomography application is part of our EM control software that handles the setting of the EM to various modes of operation, and includes the custom magnification steps needed for imaging with the energy filter.

Microanalysis

The STEM capability of the IVEM is particularly useful for microanalysis. An annular darkfield STEM detector allows simultaneous STEM imaging and spectroscopy. By means of Gatan EELS software, and our 4pi Revolution EDX/STEM imaging system, EELS and EDX spectra can be recorded simultaneously at each pixel in an image. For quantitative analysis of trace elements such as calcium and phosphorus, a spectrum can be acquired from an area selected in STEM mode, and analysis can be carried out using Gatan EELS analysis software. Elemental maps can also be made in EFTEM mode. Our EM control software makes it convenient to switch between the several microanalytical modes.

Phase plate imaging

The IVEM is equipped with a Zernike phase plate, intended for high-contrast in-focus imaging of frozen-hydrated specimens. Integration of phase-plate imaging with low-dose tilt-series collection is under development.

Tour of the IVEM

Tour of the IVEM

A: Keyboard for stage computer
B: Monitor for TVIPS application
C: Monitor energy-filtered imaging application
D: Monitor for EM control
E: Dewar for EDX detector
F: SEM detector
G: STEM/SEM unit (ASID)

H: TV for energy filter alignment
I: Computer for energy filter
J: Workstation for image post-processing

Inset:
equipment behind kneespace panel K: TVIPS FastScan camera
L: STEM detector
M: Energy filter