How to use the microscope to observe with the cutting method
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If a microscope sample uses styrene resin as the embedding medium, it's quite convenient to perform scanning electron microscopy (SEM) observations by sectioning. Since styrene is a polymer, solvents like propylene oxide and isovaleric acid can easily dissolve it. By changing the solvent several times, the styrene can be effectively removed. Therefore, when preparing TEM specimens, multiple embedded capsules are typically made. After the sample is polymerized, it is cut using the previously described method. The McAudi microscope then places the cut sample in propylene oxide for about two hours, followed by replacement with isocyanuric acid and critical point drying.
In addition to the resin-based cutting method, there are other techniques such as organic solvent-based methods (e.g., alcohol or isovaleric acid), water-soluble embedding agents (e.g., dimethyl hydrazine or glycerin), and alternative approaches like freezing or freeze-fracturing. These methods are generally easier to perform. For example, with the alcohol-based cutting method, the sample is first fixed and dehydrated using alcohol. Once dehydration is complete, the sample is placed into a capsule containing anhydrous ether, ensuring no bubbles form. The capsule is then placed in a low-temperature environment to allow the resin to cure. Afterward, the sample is sectioned. The cut sample is immediately immersed in anhydrous ethyl ether at room temperature, where it melts and is then transferred to isoamyl acetate for critical point drying.
It’s important to note that specimens prepared using the cutting method require observation under a high-resolution microscope, such as a field emission scanning electron microscope. Conventional SEMs have lower resolution compared to transmission electron microscopes (TEMs). If you're observing cellular structures or tissues, a field emission gun is necessary to achieve the clarity needed for detailed imaging. Due to the strong three-dimensional effect of SEM, cut samples reveal the spatial arrangement of structures like the Golgi apparatus, large and small vacuoles, and mitochondria, which appear as spacers or stretched filaments. The nucleus may look like a small mushroom attached to the "plate" of the endoplasmic reticulum. However, some researchers argue that polarized light microscopy doesn't provide new insights into cell structure, which is why this method isn’t widely adopted.