Transmission electron microscopy 214513 221470747 2008-06-24T17:19:58Z 212.20.74.230 [[cs:Transmisní elektronový mikroskop]] {{redirect|TEM}} [[Image:TEM.jpg|thumb|right|A section of a cell of ''[[Bacillus subtilis]]'', taken with a Tecnai T-12 TEM. The scale bar is 200nm.]] '''Transmission electron microscopy''' ('''TEM''') is a [[microscope|microscopy]] technique whereby a beam of [[electron]]s is transmitted through an ultra thin specimen, interacting with the specimen as it passes through it. An image is formed from the electrons transmitted through the specimen, magnified and [[Focus (optics)|focused]] by an objective lens and appears on an imaging screen, a fluorescent screen in most TEMs, plus a monitor, or on a layer of [[photographic film]], or to be detected by a sensor such as a [[CCD camera]]. The first practical transmission electron microscope was built by Albert Prebus and [[James Hillier]] at the [[University of Toronto]] in 1938 using concepts developed earlier by [[Max Knoll]] and [[Ernst Ruska]]. ==Background== [[Image:Elektronenmikroskop.jpg|100px|thumb|right|An old model TEM, the HV source begins at the top, and the electromagnetic lenses focus the electrons from the HV source through the specimen and onto the viewing screen (bottom) A photographic emulsion can be loaded into the drawer at the base]] Theoretically the maximum resolution that one can obtain with a light microscope has been limited by the [[wavelength]] of the [[photons]] that are being used to probe the sample and the [[numerical aperture]] of the system. Early twentieth century scientists theorized ways of getting around the limitations of the relatively large wavelength of [[Visible spectrum|visible light]] (wavelengths of 400–700 [[nanometer]]s) by using electrons. Like all matter, electrons have both wave and particle properties (as theorized by [[Louis-Victor de Broglie]]), and their wave-like properties mean that a beam of electrons can be made to behave like a beam of electromagnetic radiation. Electrons are usually generated in an electron microscope by a process known as [[thermionic emission]] from a filament, usually tungsten, in the same manner as a [[light bulb]], or by [[field emission]]. The electrons are then accelerated by an [[electric potential]] (measured in V, or [[volts]]) and focused by electrostatic and electromagnetic lenses onto the sample. The transmitted beam contains information about electron density,[[Phase (waves)|phase]] and periodicity; this is used to form an image. ===Contrast Formation=== Details in a light microscope sample can be enhanced by [[staining|stains]] that absorb light. TEM stains enhance contrast by scattering electrons out of the beam. Compounds of [[heavy metals]] such as [[osmium]], [[lead]], or [[uranium]] selectively deposit electron dense atoms in or on the sample. Beam electrons are deflected by electrostatic interactions with the positive atomic nuclei and negative electron clouds. The scattered electrons are excluded from the transmitted beam by the objective aperture. Electron dense areas in the sample appear darker on the screen and on positive images. ==Modifications== The capabilities of the TEM can be further extended by additional stages and detectors, sometimes incorporated on the same microscope. An ''electron cryomicroscope'' is a TEM with a specimen holder capable of maintaining the specimen at [[liquid nitrogen]] or [[liquid helium]] temperatures. This allows imaging specimens prepared in [[vitreous ice]], the preferred preparation technique for imaging individual molecules or macromolecular assemblies. A TEM can be modified into a [[scanning transmission electron microscope]] (STEM) by the addition of a system that rasters the beam across the sample to form the image, combined with suitable detectors. An analytical TEM is one equipped with detectors that can determine the elemental composition of the specimen by analysing its [[X-ray]] spectrum or the energy-loss spectrum of the transmitted electrons. Modern research TEMs may include [[aberration]] correctors, to reduce the amount of distortion in the image, allowing information on features on the scale of 0.1&nbsp;nm to be obtained (resolutions down to 0.05&nbsp;nm have been achieved <ref>[http://www.physorg.com/news108313701.html TEAM Project Achieves Microscopy Breakthrough]</ref>) at magnifications of 50 million times <ref>[http://www.sc.doe.gov/bes/scale_of_things.html The Scale of Things (Office of Basic Energy Sciences)]</ref>. [[Monochromator]]s may also be used which reduce the energy spread of the incident electron beam to less than 0.15&nbsp;[[electronvolt|eV]]. Major TEM makers include [[JEOL]], [[Hitachi High-technologies]], [[FEI Company]] (from merging with [[Philips]] Electron Optics) and [[Carl Zeiss]]. ==Applications of the TEM== The TEM is used heavily in both [[material science]]/[[metallurgy]] and the [[biological sciences]]. In both cases the specimens must be very thin and able to withstand the high vacuum present inside the instrument. For biological specimens, the maximum specimen thickness is roughly 1 micrometre. To withstand the instrument [[vacuum]], biological specimens are typically held at [[liquid nitrogen]] temperatures after embedding in vitreous ice, or fixated using a [[negative stain]]ing material such as [[uranyl acetate]] or by plastic embedding. Typical biological applications include [[tomogram|tomographic]] reconstructions of small cells or thin sections of larger cells and 3-D reconstructions of individual molecules via [[Single Particle Reconstruction]]. [[Image:Fib_tem_sample.jpg|thumb|right|[[Scanning electron microscope|SEM]] image of a thin TEM sample milled by [[focused ion beam|FIB]]. The thin membrane shown here is suitable for TEM examination; however, at ~300-nm thick, it would not be suitable for high-resolution TEM without further milling.]] In material science/metallurgy the specimens tend to be naturally resistant to vacuum, but must be prepared as a thin foil, or etched so some portion of the specimen is thin enough for the beam to penetrate. Preparation techniques to obtain an electron transparent region include [[ion beam]] milling and wedge polishing. The [[focused ion beam]] (FIB) is a relatively new technique to prepare thin samples for TEM examination from larger specimens. Because the FIB can be used to micro-machine samples very precisely, it is possible to mill very thin membranes from a specific area of a sample, such as a semiconductor or metal. Materials that have dimensions small enough to be electron transparent, such as powders or nanotubes, can be quickly produced by the deposition of a dilute sample containing the specimen onto support grids. The suspension is normally a volatile solvent, such as ethanol, ensuring that the solvent rapidly evaporates allowing a sample that can be rapidly analysed. The imaging techniques explained below are particularly important in materials science. Faults in crystals affect both the mechanical and the electronic properties of materials, so understanding how they behave gives a powerful insight. By carefully selecting the orientation of the sample, it is possible not just to determine the position of defects but also to determine the type of defect present. If the sample is orientated so that one particular plane is only slightly tilted away from the strongest diffracting angle (known as the [[Bragg diffraction|Bragg Angle]]), any distortion of the crystal plane that locally tilts the plane to the Bragg angle will produce particularly strong contrast variations. However, defects that produce only displacement of atoms that do not tilt the crystal to the Bragg angle (i.e. displacements parallel to the crystal plane) will not produce strong contrast. Furthermore, the [[HRTEM]] technique (see below) allows the direct observation of crystal structure and therefore has an advantage over other methods in that there is no displacement between the location of a defect and the contrast variation caused in the image. However, it is not always possible to interpret the lattice images directly in terms of sample structure or composition. This is because the image is sensitive to a number of factors (specimen thickness and orientation, objective lens defocus, spherical and chromatic aberration), and although quantitative interpretation of the contrast shown in lattice images is possible, it is inherently complicated and may require extensive simulation of the images. Computer modeling of these images has added a new layer of understanding to the study of crystalline materials. ==Imaging in the TEM== [[Image:TEM1.jpg|right|thumb|230px|Transmission Electron Micrograph of [[Dislocation]]s]] The contrast in a TEM image is not like the contrast in a light microscope image. A crystalline material interacts with the electron beam mostly by [[diffraction]] rather than [[Absorption (electromagnetic radiation)|absorption]], although the intensity of the transmitted beam is still affected by the volume and density of the material through which it passes. The intensity of the diffraction depends on the orientation of the planes of atoms in a [[crystal]] relative to the electron beam; at certain angles the electron beam is diffracted strongly from the axis of the incoming beam, while at other angles the beam is largely transmitted. Modern TEMs are often equipped with specimen holders that allow the user to tilt the specimen to a range of angles in order to obtain specific diffraction conditions, and apertures placed below the specimen allow the user to select electrons diffracted in a particular direction. A high-contrast image can therefore be formed by blocking electrons deflected away from the optical axis of the microscope by placing the aperture to allow only unscattered electrons through. This produces a variation in the electron intensity that reveals information on the [[crystal structure]], and can be viewed on a [[fluorescent]] screen, or recorded on [[photographic film]] or captured electronically. [[Image:char_tem.jpg|right|thumb|230px|Transmission Electron Micrograph of a [[cobalt]] [[catalyst]] (darker spots) supported on [[coal]] carbonized at 850 ºC and added by ion-exchange technique]] This technique (known as ''Bright Field'' or ''Light Field'') is particularly sensitive to extended crystal lattice defects in an otherwise ordered crystal, such as [[Dislocation|dislocations]]. As the local distortion of the crystal around the defect changes the angle of the crystal plane, the intensity of the scattering will vary around the defect. As the image is formed by the distortion of the crystal planes around the defect, the contrast in these images does not normally coincide exactly with the defect, but is slightly to one side. It is also possible to produce an image from electrons deflected by a particular crystal plane. By either moving the aperture to the position of the deflected electrons, or tilting the electron beam so that the deflected electrons pass through the centred aperture, an image can be formed of only deflected electrons, known as a ''Dark Field'' image. In the most powerful diffraction contrast TEM instruments, crystal structure can also be investigated by [[High Resolution Transmission Electron Microscopy]] (HRTEM), also known as phase contrast imaging as the images are formed due to differences in phase of electron waves scattered through a thin specimen. Resolution of the HRTEM is limited by [[Lens (optics)#Spherical_aberration|spherical]] and [[Lens (optics)#Chromatic_aberration|chromatic aberration]], but a new generation of aberration correctors has been able to overcome spherical aberration. Software correction of spherical aberration has allowed the production of images with sufficient resolution to show carbon atoms in diamond separated by only 0.89 [[ångström]]s (89 [[picometer|pm]], one ångström is 0.0000000001 of a meter or 100 picometers) and atoms in silicon at 0.78 ångströms (78 pm) at magnifications of 50 million times. Improved resolution has also allowed the imaging of lighter atoms that scatter electrons less efficiently &mdash; lithium atoms have been imaged in lithium battery materials<ref>[http://www.fei.com/Portals/_default/PDFs/content/2006_06_LithiumImagingOkeefe_wp.pdf Imaging lithium atoms at sub-Ångström resolution] pdf, 6Mb </ref>. The ability to determine the positions of atoms within materials has made the HRTEM an indispensable tool for [[nanotechnology]] research and development in many fields, including [[heterogeneous catalysis]] and the development of [[semiconductor device]]s for electronics and photonics. ==Limitations== There are a number of drawbacks to the TEM technique. Many materials require extensive sample preparation to produce a sample thin enough to be electron transparent, which makes TEM analysis a relatively time consuming process with a low throughput of samples. The structure of the sample may also be changed during the preparation process. Also the field of view is relatively small, raising the possibility that the region analysed may not be characteristic of the whole sample. There is potential that the sample may be damaged by the electron beam, particularly in the case of biological materials. ==See also== {{wikibooks|Nanotechnology|Electron_microscopy#Transmission_electron_microscopy_.28TEM.29|Transmission electron microscopy (TEM)}} *[[Scanning electron microscope]] *[[Electron microscope]] *[[Transmission Electron Aberration-corrected Microscope]] *[[Energy filtered transmission electron microscopy]] *[[Electron diffraction]] ==References== <references/> ==External links== *http://esteem.ua.ac.be/ *[http://ncem.lbl.gov The National Center for Electron Microscopy, Berkeley California USA] *[http://ncmi.bcm.tmc.edu The National Center for Macromolecular Imaging, Houston Texas USA] *[http://nramm.scripps.edu/ The National Resource for Automated Molecular Microscopy, La Jolla California USA] *http://www.unl.edu/CMRAcfem/em.htm *http://www.matter.org.uk/tem/ *http://www.microscopy.ethz.ch *[http://www.fei.com/ Manufacturer of TEM's (FEI Company)] *[http://www.lv-em.com Delong Group] *[http://www.rodenburg.org/ Tutorial courses in Transmission Electron Microscopy] *[http://www.msm.cam.ac.uk/doitpoms/tlplib/tem/index.php Cambridge University Teaching and Learning Package on TEM] [[Category:Microscopes]] [[Category:Microscopy]] [[Category:Scientific techniques]] [[cs:Transmisní elektronový mikroskop]] [[de:Transmissionselektronenmikroskop]] [[es:Microscopio electrónico de transmisión]] [[fa:میکروسکوپ الکترونی عبوری]] [[fr:Microscopie électronique en transmission]] [[hu:Transzmissziós elektronmikroszkóp]] [[ja:透過型電子顕微鏡]] [[vi:Kính hiển vi điện tử truyền qua]] [[tr:Geçirimli elektron mikroskobu]] [[zh:透射电子显微镜]]