Electron microscopes are very powerful tools for visualizing biological samples. They allow scientists to look at cells, tissues and small organisms in great detail. With electron microscopes, scientists and researchers can see the micron and nanoscale world around us.
The two most common types of electron microscopes are transmission electron microscopy (TEM) and scanning electron microscopy (SEM). In general, the SEM is ideal if you need to see a relatively large area and only need surface detail. If you need internal details of a small sample with near atomic resolution, the TEM must be used.
To observe these biological samples, one must go through a preparation step to help the sample withstand the environment inside the microscope. Creative Biolabs describes a common protocol for electron microscopy, including the process of preparing specimens of cells, tissues and other samples, to follow standard procedures for life science analysis with the aid of a microscope.
|Solutions and Reagents
|Phosphate buffer (PBS), aldehyde fixative, fixative buffer, washing buffer, ethanol solution
Electron microscopes can excite electrons to form images, providing amazing levels of fine magnification. Here, we describe the general operation of electron microscopy using SEM as an example for reference. Please note that we recommend discussing this with our staff prior to starting your electron microscopy protocol.
There are many variations of fixation procedures depending on the type of sample to be examined. First choose the appropriate aldehyde fixative to fix the sample for at least one hour. For tissue samples, this can be done after initial fixation, rinsing in buffer and post-fixing in osmium tetroxide for at least 1 hour. After fixation, rinse in buffer.
After rinsing in buffer, dehydrate the samples through a series of graded 50-100% alcohols. After dehydration, perform drying. You can choose between critical point drying (CPD) or chemical drying.
Mount the sample on a specimen holder and sputter coat it with Au/Pd. The SEM sample is coated with a thin layer of metal (usually gold or gold-palladium). The metal coating makes the sample electrically conductive and draws electrons away from the bombarded sample.
Image and observe gold-plated samples with a scanning electron microscope. The imaging is performed under high vacuum and accelerating voltage.
The electron microscope is a precision instrument, and attention must be paid to sample preparation before observing the sample. If there is a problem with the sample or improper preparation, not only can we not get the desired effect, but also will cause damage to the electron microscope and affect the performance. Therefore, you should pay attention to the following issues when preparing samples.
The samples need to be dry and free of water and volatile solvents. If the sample contains water or volatile solvents, they may evaporate in the vacuum environment causing the electron microscope filament to malfunction.
Poor electrical conductivity
Poor thermal stability
After checking the reasonable troubleshooting steps, if you still have problems, you can contact us for inquiries.
For research use only. Not intended for any clinical use.