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Electron Microscopy Sciences

EMS Academy Applications

arrow13The Cryo SEM Workshop

This course will cover the process of rapid freezing, fracturing, coating and imaging of a variety of samples.

Examples of the endless possibilities when doing Cryo SEM

Vertebrates - Tissues


Fractures through the myocardium
Small pieces of heart have been mixed, washed in buffer then cryo-protected with 30% glycerol. Contracted myofibrils can be seen in longitudinal aspect. The fracture in places has occurred along the sarcolemma and the positions of the Z-lines (marked by 'z') and the T-tubules (marked by arrows) are clearly visible in places. The intimate relationship between the capillaries and the myofibrils can be appreciated from this preparation. The contents of the capillary have a smooth appearance due to the presence of glycerol. Bar: 5µm

 
Frozen hydrated mouse lung
Low-temperature SEM retains all of the cellular and extra-cellular fluids in the lung tissue. Consequently, the micrograph clearly illustrates that the bronchioles are covered with a thin layer of mucus (marked by arrows). None of the alveoli show any collapse. Bar: 5µm

Fungi

 

Carnivorous Plants


Fungal spore sac
Spore sac of a fungus. The sacs tend to burst open as soon as they are exposed to moisture, so the only way to visualise them is to use cryo-SEM. Image courtesy of Miranda Waldron at the Electron Microscope Unit at the University of Cape Town, South Africa

 
Sundew leaf and insect
A small fly trapped on a carnivorous South African sundew plant. The whole plant is about 3cm across and has sticky hairs on its leaves to capture small insects. Image courtesy of Miranda Waldron at the Electron Microscope Unit at the University of Cape Town, South Africa

Foodstuffs – Various


Mayonaisse – fractured
Image courtesy of FEI Company

 
Chocolate Bar

Targeted Participants

Individuals who are new to the field of cryo SEM or desire a technical refresh to maintain current skills or just those that want to see and learn all of the possibilities of the technology.

Facility

The EMS Microscopy Academy
Located in Hatfield, Pennsylvania, the Academy provides electron microscopy classes, workshops and training sessions for all fields of microscopy, including materials science and biological science.

Scope of Class

Many types of samples such as ice cream, pastes, paints, and gels do not lend themselves to routine SEM sample preparation methods such as critical point drying or freeze drying due to the morphologic changes caused by desiccation. To avoid these artifacts it is necessary to image the hydrated or natural state. This requires that the sample be rapidly frozen, to reduce ice crystal damage, and fractured to reveal their physical / functional sub-surface morphology. These samples are applied to the stub, immersed in liquid nitrogen slush loaded into the Cryo fracturing and coating chamber, and finally the SEM. This final preparation takes place in a high vacuum environment thus minimizing the possibility of frost contamination. Within the SEM, while viewing, the sample temperature can be maintained at -130° or warmed slightly to facilitate sublimation of the surface. The selection of accelerating voltage (kV), for surface detail, and spot size for resolution and charging are critical and will be covered in detail.

Parameters such as working distance which affects depth of field (Dfi) and resolution, plus tilt and raster rotate will be examined for proper image collection.

Format

Lecture, demonstration and hands-on practice, as well as round table discussion. Participants are encouraged to bring their own samples, if possible.

Main Curriculum

  • Theory and overview of cryo SEM
  • Mounting and adhering
  • Freezing, loading and fracturing
  • Insitu coating
  • Operation of SEM
  • Cryo face-off Leica UC 7 Crion
  • Specific techniques of Cryo SEM imaging

Equipment

Hitachi S3500 SEM PP3010T Cryo Preparation System
Leica UC7 Ultramicrotome Leica Crion Cryomicrotome

Faculty

Al Coritz
Al has been doing Electron Microscopy for 38 years, beginning at the Yale School of Medicine and ending up on the commercial side with several key EM companies. His specialty is Cryo-techniques and Thin Film Technology: i.e. Freeze Fracture/Rotary Shadowing, High Pressure Freezing, and more. He is currently with Electron Microscopy Sciences where he has been the Technical Director for over 20 years.

Michael Kostrna
Michael was the program director of the Electron Microscopy Technician program at Madison Area Technical College and has more than 35 years in EM technical education and research experience. He has been training EM students for 29 years and has developed curricula and lab exercises for TEM, SEM, OLM, lab safety, introductory and advanced biological EM, EM, maintenance, and x-Ray microanalysis. He has worked with companies such as SC Johnson Polymer, Dow Chemicals, Io Genetics, Virent Technologies, ABS Global, NanoOnocology, and Microscopy Inovations, and in the process gained insight to the various applications of EM.

View our schedule to see if this course is currently offered.

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