EMCF Equipment & Methods

FEI Tecnai F20 TEM

FEI Tecnai F20 TEM, equipped with a field emission gun and bottom mounted 4K camera is located in Room U017 at the Bioquant. This 200KV transmission electron microscope can be controlled by the SerialEM software to collect data for EM tomography which is further processed in eTomo to generate tomograms and IMOD is used for making 3D models. This technique is termed Electron tomography and allows observation of the ultrastructure in 3D at high resolution. This is achieved with an electron microscope that uses a higher acceleration voltage (200kV and higher) to penetrate thicker (~ 200-500 nm) sections. By taking images of the section at a series of tilt angles, a reconstruction of the 3 dimensional structures within the section can be achieved. Consecutive sections can then be used to reconstruct a larger volume.

JEOL JEM1400 TEM

JEOL JEM1400 transmission electron microscope (TEM) is available at INF345 R027. The microscope’s original camera has been replaced with a bottom mount 4k x 4k pixel digital camera (TemCam F416; TVIPS, Gauting, Germany). The TemCam F416 allows stitching of individual small images to obtain overviews at high resolution. In addition to EM-Menu, SerialEM can be used to acquire images. On Nov 2023 the old tungsten filament on the JEOL was upgraded to a Lanthanum Hexaboride (LaB6) filament for a stronger beam with higher longevity and better resolution.

Zeiss Leo 1530 SEM

Zeiss Leo 1530 SEM is available at INF 307, Room U06. It is a high resolution scanning EM with a Schottky field emission gun, an SE2 and an in-lens detector. The in-lens detector yields a good image contrast at low beam currents. Its acceleration voltage range from 200V to 30kV. As well as the conventional topographical imaging, the SEM can be used for imaging ultrathin sections. For example, for array tomography (AT).

BAL-TEC HPM 010 High Pressure Freezer 

High-Pressure Freezing at 2100 bar pressure and liquid nitrogen temperatures drastically reduces ice crystal formation inside specimen. This allows physical fixation in which amorphous freezing of cell fluids occurs. High pressure freezing also increases chances to preserve antigenicity and fluorescent signal through the sample preparation. High pressure freezing is therefore important for immunolabelling and correlative light and electron microscopy techniques. However, specimen size for high pressure freezing has to be very small to achieve good freezing. Material has to be fitted into planchets which are usually up to 200 µm in depth.

Leica AFS II 

We have three Leica Automatic Freeze Substitution units with two EM FSP freeze substitution Automated robots. 

During Freeze substitution the ice is substituted against a liquid solvent at low temperature which is in turn substituted for a resin. Fixatives or contrasting chemicals can be added to the freeze substitution medium as needed. The equipment for freeze substitution and low temperature embedding is located very conveniently in the lab next door to the high pressure freezer inside a special fume-hood safety cabinet. The Leica AFS units are equipped with a binocular microscope and exchange of freeze substitution media can be done manually or with the help of a robot.

Bruker SkyScan 1272 microCT

An X-ray tomography system for visualization and measurement of complete three-dimensional object structures, after fixation and staining . The tomograms reaches a spatial resolution in the level of 1 micron. The internal structure can be non-destructively reconstructed and fully analyzed. The X-ray scanner contains a microfocus X-ray source, an X-ray detector, an object manipulator for positioning and rotation of the objects during tomographic acquisition. It has a Microfocus sealed X-ray source 20-100kV, max. 10W, <5um spot size @ 4W, an X-ray detector, and 14bit cooled CCD fiberoptically coupled to scintillator.

Leica EM UC7 Ultramicrotome 

We have 3 Leica EM UC7, a Leica EM FC6, a Leica Ultracut UCT, and a Reichert Ultracut S for semi- and ultra-thin sections. Also, we now offer an Array Tomography (AT) service for 3D reconstruction of small to medium size volumes using SEM. This is a relatively new setup for serial sectioning and mounting samples on coverslips, using one of our UC7 Ultramicrotomes. We aim to facilitate understanding and answers to some of the more complex questions about cellular morphology in 3D.

PELCO BioWave® Pro+ 

The laboratory microwave oven tissue processor further enhances processing of tissue for electron microscopy, histology, immunofluorescence and confocal microscopy better and faster than conventional methods. It has a continuous microwave power setting range from 100-750 Watts. Microwave frequency of 2.45 Ghz, power control, programmable controller with 10 modifiable presets, 6" touch screen user interface, access to custom protocols, and temperature control of ± 1°C for most aqueous solutions.

Leica CPD300 

The replacement of organic solvent with a transitional fluid as liquid carbon dioxide is done with a Critical Point Dryer at EMCF. This technique prevents collapse and shrinkage of the specimen caused by air-drying. Conductivity is conferred afterwards by gold sputter coating.

Leica ACE600 Carbon Coater and Sputter Coater 

Precisely applying a thin layer of carbon is for example used to increase stability of the film on grids used for mounting sections. This can be done using the carbon coater mode of ACE600. The ACE600 also has a sputter coater mode which is used for coating with metal. Available targets are Gold-Palladium, Gold and Platinum/Palladium. For conventional imaging in the SEM, specimen surface must be conductive and electrically grounded. Hard, dry material can be examined in an SEM with little further treatment; however, cells, tissues and soft-bodied organisms require chemical fixation and dehydration, critical point drying, followed by sputter coating.