Sample Preparation Techniques
For instructions on how to coat samples, etc, download the relevant files
QUANTOMIX - wet sample preparation
Until recently, you could only view inanimate, dry materials with the SEM. However this has changed, as a company called Quantomix has developed and commercialized breakthrough solutions based on its proprietary WETSEM-Technology. This technology enables scanning electron microscopes to image and analyze wet samples such as cells, tissue biopsies, foods and ink, in their native environment. Eliminating the need for time-consuming preparation procedures, the WETSEM-Technology ensures that sample integrity is not compromised by artifacts.
Examples of wet samples using Quantomix Click images to enlarge:
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A cell which has taken in particles. |
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Primary fibrolost with PTA staining. |
In order to be able to take photos like these, 15 μl of the solution is placed inside the capsule (see picture, right), which is then viewed under the microscope. Quantomix capsules [1] rely on a thin electron-transparent membrane that enables separation, and thereby protection, of the wet sample from the electron microscope vacuum (fig. 1). Imaging is performed using a standard SEM combined with a Back Scattered Electron detector. There are also other general advantages of using Quantomix capsules other than its most obvious benefit: the ability to image wet samples in an EM. A significant advantage of quantomix over other electron microscopy techniques is the ability to image samples a few millimeters thick without any time-consuming, costly processing such as thin sectioning, embedding, freezing or coating.
Figure 1: A cut-through representation of a Quantomix capsule
FREEZE DRYING
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Freeze drying (also known as Lyophilization) is a dehydration process typically used to preserve a perishable material. Freeze drying works by freezing the material and then reducing the surrounding pressure to allow the frozen water in the material to sublime directly from the solid phase to gas. The application of high vacuum in freeze drying sublimes ice much more quickly, making it useful as a deliberate drying process. A cold condenser chamber and/or condenser plates provide a surface(s) for the vapour to re-solidify on. These surfaces must be colder than the temperature of the surface of the material being dried, or the vapour will not migrate to the collector. Temperatures for this ice collection are typically below -50 °C. For more information, click here (pdf). |
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CRITICAL POINT DRYING
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During the phase change from liquid to gas, the surface tension that occurs between the gas-liquid interface can cause damage. The surface tension is avoided by drying with a solution that is taken from subcritical to supercritical, which avoids a gas-liquid interface because the densities of the gas and liquid are equivalent. Water is not used because its critical point is very high, so the water is displaced with alcohol or acetone. More than one substitution may have to be made because the solution that displaces the water must be miscible with the supercritical solution. After dehydration, the specimen is placed in a subcritical solution. Carbon dioxide is the most frequently used substance. The temperature and pressure are increased until the solution is supercritical and the liquid changes to gas without damaging the sample. For more information, click here (.pdf). |
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1. Ruach-NirAn, I. 'Innovative Method for Imaging and Chemical Analysis of Wet Samples in Scanning Electron Microscopes', Microscopy Today, July 2005, 10-14