Transmission+Electron+Microscopy

__**Transmission Electron Microscopy **__


 * Basic Description: **

Transmission Electron Microscopy (TEM) is a technique used to show the fine structures of a cell that can't be seen with visible light. TEM is similar to a light microscope except for the fact that it's larger and "upside down" with a 200 times better resolution. TEMs work by shooting electrons completely through the sample in order to create an image the observer can look at. How this works is by using a tungsten filament to produce an electron beam in a vacuum chamber. The electrons that are emitted are accelerated through an electromagnetic field which narrowly focuses the electron beam through the material the observer is sampling. The electrons that pass through hit a phosphor screen, on a layer of photographic film, or to be detected by a sensor such as a CCD camera producing the image for the observer. Where the sample has less “stuff” in the way (lower density) more electrons are able to pass through creating a brighter image. Therefore the opposite is true where the more density there is the darker the image is produced due to less electrons being able to pass through.[1]


 * Purpose of Technique: **

The purpose of this technique is to see the different components inside the membrane rather than just seeing the outside of the membrane by creating an image at a very high resolution due to the smaller wavelength electrons travel on (about 100,000 times shorter than visible light). It is used to reveal the shape of individual macromolecules at a very high resolution of around 0.002 nm.[1] TEM produces a major analysis method in physical and biological sciences including research for cancer, virology and pollution.




 * Origin and History: **

The origins of the technique came originally about by Ernst Abbe who stated that resolution of an image is dependent on the wavelength of the light. In 1858, Plücker discovered that the deflection of electrons (known as cathode rays) was possible by utilizing magnetic fields, which was tested by Braun in 1898 to build primitive cathode ray oscilloscopes (CROs). It was discovered in 1891 by Riecke that cathode rays could be focused by these magnetic fields with simple lens designs, which was more understood by Busch in 1926 that showed that the lens maker’s equation could be applicable to electrons. After all these discoveries, the 1st TEM was built by Max Knoll’s and a group of researchers from the Technological University of Berlin in 1931. It was also the 1st group to develop a TEM with resolving power greater than the resolving power of light (1933) and production of the 1st commercial TEM in 1939.


 * Recent Research: **

There are many articles that utilize this technique. There was a study on the actin cytoskeleton of whole-mount breast cancer cells using TEM at different resolutions. The TEM images revealed the positions of intermediate electron dense DRMs. TEM also showed the change of the complex organization of actin cytoskeleton structures in more detail when in interaction with EGF since the response time of EGF with MTLn3 cells is changed within minutes.[2] Another article uses TEM for virus detection. Using TEM at 18000 magnification, supernatants from epithelioma papulosum cyprinicell culture with the cytopathic effect. At this magnification, TEM showed the presence of iridovirus-like particles which were then tested using PCR methods for ranavirus detection.[4] TEM is used for characterizing atmospheric aerosol particles. When quantitative single-particle analysis is applied with TEM-EDX, it is shown that the Monte Carlo calculation method can be used which shows that the technique is functional and dependable for the characterization of submicron aerosol particles. This can also be shown using TEM coupled with energy-dispersive X-ray spectrometry.[3]

Another research article that utilizes the TEM technique is “Rapid determination of the toxicity of quantum dots (QDs) with luminous bacteria” In this paper, the authors were trying to evaluate the toxicity of quantum dots by measuring the decrease of the light emitted by luminous bacteria (a bacteria that are a well-known species that emit bright bioluminescence which is directly proportional to the metabolic activity of the bacterial population). This was observed by using a transmission electron microscope to determine how the QDs reacted with the bacteria. The results showed that after the QDs were made present they indeed accumulated inside the bacteria. As they entered the bacteria they changed from a size of 3-4 nm to 20-30 nm. This showed that the QDs, if present, could indeed effect the function of the bacteria and was shown with TEM imaging. Another paper that utilized TEM imaging is “Elemental distribution analysis of type I collagen fibrils in tilapia fish scale with energy-filtered transmission electron microscope” the purpose of this experiment was to discover the elemental distribution of calcium, phosphorus, oxygen, and carbon in a single collagen fibril obtained from tilapia fish scales. After analyzing the tilapia scale through TEM it was clear that there were dark and bright areas of the cell. The dark area observed were attributed to the presence of heavier metals like calcium and phosphorus while the lighter areas was carbon. Due to the location of the specific elements in the cell (discovered through TEM) it is suggested that the mineralization of collagen fibrils depends on the elements present in the internal layer.

A last article that uses TEM as a technique is “Abnormal elongation of midpiece, absence of axoneme and outer dense fibers at principal piece level, supernumerary microtubules: a sperm defect of possible genetic origin” In this experiment, researches analyzed a 42 year old infertile man sperm and compared them to known fertile sperm. It was shown with while using TEM that the sperm of the infertile male had an abnormal length extension in the longitudinal section of the midpiece region compared to the fertile sperm.


 * References: **

Alberts, Bruce et. al. (2008). The Molecular Biology of the Cell. Ed 5. Garland Science, Taylor & Francis Group, LLC, New York, NY.

Jahn, K.A. et. al. (2009). “Correlative fluorescence and transmission electron microscopy:an elegant tool to study the actin cytoskeleton of whole-mount(breast) cancer cells.” Journal of Microscopy. 235(3): 282–292. doi: 10.1111/j.1365-2818.2009.03223.

Manoj Kumar Singh, B. N. (2012). The evolution of Cefotaximase Activity in the TEM B-Lactamase. Journal of molecular biology, 205-220.

<span style="font-family: 'Times New Roman',Times,serif; font-size: 140%;">Maskey, Shila & Ro, Chul-un. (2011). "Quantitative energy-dispersive electron probe X-ray microanalysis for single-particle analysis and its application for characterizing atmospheric aerosol particles." Indian Academy of Sciences. 76(2): 281-292.

<span style="font-family: 'Times New Roman',Times,serif; font-size: 140%;">Ruska, E. (2008, January 3rd). The Transmission Electron Microscope. Retrieved MArch 4, 2012, from Nobelprize.org: http://www.nobelprize.org/educational/physics/microscopes/tem/index.html

<span style="font-family: 'Times New Roman',Times,serif; font-size: 140%;">Vesely, T. et. al. (2011) “Investigation of ornamental fish entering the EU for the presence of ranaviruses.” Journal of Fish Diseases. 34: 159–166. doi:10.1111/j.1365-2761.2010.01224.