Preparing Specimens for Electron Microscopy

Advertisements
Advertisements

Related Posts:


(a) An ultramicrotome used to prepare specimens for a TEM. (b) A technician uses an ultramicrotome to slice a specimen into thin sections. (credit a: modification of work by “Frost Museum”/Flickr; credit b: modification of work by U.S. Fish and Wildlife Service Northeast Region)

OpenStax Microbiology

Samples to be analyzed using a TEM must have very thin sections. But cells are too soft to cut thinly, even with diamond knives. To cut cells without damage, the cells must be embedded in plastic resin and then dehydrated through a series of soaks in ethanol solutions (50%, 60%, 70%, and so on). The ethanol replaces the water in the cells, and the resin dissolves in ethanol and enters the cell, where it solidifies. Next, thin sections are cut using a specialized device called an ultramicrotome. Finally, samples are fixed to fine copper wire or carbon-fiber grids and stained—not with colored dyes, but with substances like uranyl acetate or osmium tetroxide, which contain electrondense heavy metal atoms.

When samples are prepared for viewing using an SEM, they must also be dehydrated using an ethanol series. However, they must be even drier than is necessary for a TEM. Critical point drying with inert liquid carbon dioxide under pressure is used to displace the water from the specimen. After drying, the specimens are sputter-coated with metal by knocking atoms off of a palladium target, with energetic particles. Sputter-coating prevents specimens from becoming charged by the SEM’s electron beam.

Source:

Parker, N., Schneegurt, M., Thi Tu, A.-H., Forster, B. M., & Lister, P. (n.d.). Microbiology. Houston, Texas: OpenStax. Access for free at: https://openstax.org/details/books/microbiology

Advertisements
Advertisements