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53-405: Lamins were first identified in the cell nucleus, using electron-microscopy . However, they were not recognized as vital components of nuclear structural support until 1975. During this time period, investigations of rat liver nuclei revealed that lamins have an architectural relationship with chromatin and nuclear pores. Later in 1978, immunolabeling techniques revealed that lamins are localized at

106-414: A cathode-ray tube with electrostatic and magnetic deflection, demonstrating manipulation of the direction of an electron beam. Others were focusing of the electrons by an axial magnetic field by Emil Wiechert in 1899, improved oxide-coated cathodes which produced more electrons by Arthur Wehnelt in 1905 and the development of the electromagnetic lens in 1926 by Hans Busch . According to Dennis Gabor ,

159-454: A central α-helical rod domain containing heptad repeats surrounded by globular N and C-terminal domains. The N-terminal is shorter and located at the top (head) while the C-terminal is longer and located at the end (tail). Lamins have a unique structure of the heptad repeats that is continuous in nature and contains an additional six heptads. While the head domain of lamins is fairly consistent,

212-420: A lens optical system or a fibre optic light-guide to the sensor of a digital camera . Direct electron detectors have no scintillator and are directly exposed to the electron beam, which addresses some of the limitations of scintillator-coupled cameras. The resolution of TEMs is limited primarily by spherical aberration , but a new generation of hardware correctors can reduce spherical aberration to increase

265-489: A map of the angles of the electrons leaving the sample is produced. The advantages of electron diffraction over X-ray crystallography are primarily in the size of the crystals. In X-ray crystallography, crystals are commonly visible by the naked eye and are generally in the hundreds of micrometers in length. In comparison, crystals for electron diffraction must be less than a few hundred nanometers in thickness, and have no lower boundary of size. Additionally, electron diffraction

318-435: A much higher resolution of about 0.1 nm, which compares to about 200 nm for light microscopes . Electron microscope may refer to: Additional details can be found in the above links. This article contains some general information mainly about transmission electron microscopes. Many developments laid the groundwork of the electron optics used in microscopes. One significant step was the work of Hertz in 1883 who made

371-570: A much younger age. Those with HGPS typically die in their early teens, usually following a heart attack or stroke. HGPS is caused by a point mutation in the LMNA gene that codes for lamin A. The genetic alteration results in an alternative splice, creating a mutated form of prelamin A that is much shorter and lacks the cleavage site for a zinc metalloprotease. Because prelamin A cannot be properly processed during posttranslational modifications , it retains its lipid modification (farnesylation) and remains in

424-410: A sample. A few examples are outlined below, but this should not be considered an exhaustive list. The choice of workflow will be highly dependent on the application and the requirements of the corresponding scientific questions, such as resolution, volume, nature of the target molecule, etc. For example, images from light and electron microscopy of the same region of a sample can be overlaid to correlate

477-550: A single brightness value per pixel, with the results usually rendered in greyscale . However, often these images are then colourized through the use of feature-detection software, or simply by hand-editing using a graphics editor. This may be done to clarify structure or for aesthetic effect and generally does not add new information about the specimen. Electron microscopes are now frequently used in more complex workflows, with each workflow typically using multiple technologies to enable more complex and/or more quantitative analyses of

530-509: A specimen surface (SEM with secondary electrons) has also increasingly expanded into the depth of samples. An early example of these ‘ volume EM ’ workflows was simply to stack TEM images of serial sections cut through a sample. The next development was virtual reconstruction of a thick section (200-500 nm) volume by backprojection of a set of images taken at different tilt angles - TEM tomography . To acquire volume EM datasets of larger depths than TEM tomography (micrometers or millimeters in

583-641: A spectrum of heart disease ranging from no apparent effect to severe dilated cardiomyopathy leading to heart failure . Laminopathies frequently cause heart rhythm problems at an early stage in the disease process including abnormally slow heart rhythms such as sinus node dysfunction and atrioventricular block , and abnormally rapid heart rhythms such as ventricular tachycardia . As a result, those with Lamin A/C heart disease are often treated with pacemakers or implantable defibrillators in addition to medication. Electron microscope An electron microscope

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636-400: A team of researchers to advance research on electron beams and cathode-ray oscilloscopes. The team consisted of several PhD students including Ernst Ruska . In 1931, Max Knoll and Ernst Ruska successfully generated magnified images of mesh grids placed over an anode aperture. The device, a replicate of which is shown in the figure, used two magnetic lenses to achieve higher magnifications,

689-419: A working instrument. He stated in a very brief article in 1932 that Siemens had been working on this for some years before the patents were filed in 1932, claiming that his effort was parallel to the university development. He died in 1961, so similar to Max Knoll, was not eligible for a share of the 1986 Nobel prize. In the following year, 1933, Ruska and Knoll built the first electron microscope that exceeded

742-418: Is a microscope that uses a beam of electrons as a source of illumination. They use electron optics that are analogous to the glass lenses of an optical light microscope to control the electron beam, for instance focusing them to produce magnified images or electron diffraction patterns. As the wavelength of an electron can be up to 100,000 times smaller than that of visible light, electron microscopes have

795-597: Is a protein that in humans is encoded by the LMNB1 gene . The nuclear lamina consists of a two-dimensional matrix of proteins located next to the inner nuclear membrane. The lamin family of proteins make up the matrix and are highly conserved in evolution. During mitosis, the lamina matrix is reversibly disassembled as the lamin proteins are phosphorylated. Lamin proteins are thought to be involved in nuclear stability, chromatin structure, and gene expression. Vertebrate lamins consist of two types, A and B. This gene encodes one of

848-458: Is done on a TEM, which can also be used to obtain many other types of information, rather than requiring a separate instrument. Samples for electron microscopes mostly cannot be observed directly. The samples need to be prepared to stabilize the sample and enhance contrast. Preparation techniques differ vastly in respect to the sample and its specific qualities to be observed as well as the specific microscope used. To prevent charging and enhance

901-500: Is to use BSE SEM to image the block surface instead of the section, after each section has been removed. By this method, an ultramicrotome installed in an SEM chamber can increase automation of the workflow; the specimen block is loaded in the chamber and the system programmed to continuously cut and image through the sample. This is known as serial block face SEM. A related method uses focused ion beam milling instead of an ultramicrotome to remove sections. In these serial imaging methods,

954-548: The LMNA gene. Two isoforms, lamins A and C, can be created from this gene via alternative splicing . This creates a high amount of homology between the isoforms. Unlike lamin C, Lamin A is generated in a precursor form called prelamin A. Prelamin A and lamin C differ in structure only at the carboxyl-terminus. Here, prelamin A contains two extra exons that lamin C lacks. Furthermore, lamin C contains six unique amino-acid residues while prelamin A contains ninety-eight residues not found in

1007-409: The transmission electron microscope (TEM), uses a high voltage electron beam to illuminate the specimen and create an image. An electron beam is produced by an electron gun , with the electrons typically having energies in the range 20 to 400 keV, focused by electromagnetic lenses, and transmitted through the specimen. When it emerges from the specimen, the electron beam carries information about

1060-613: The 1930s, at the Washington State University by Anderson and Fitzsimmons and at the University of Toronto by Eli Franklin Burton and students Cecil Hall, James Hillier , and Albert Prebus. Siemens produced a transmission electron microscope (TEM) in 1939. Although current transmission electron microscopes are capable of two million times magnification, as scientific instruments they remain similar but with improved optics. In

1113-447: The 1940s, high-resolution electron microscopes were developed, enabling greater magnification and resolution. By 1965, Albert Crewe at the University of Chicago introduced the scanning transmission electron microscope using a field emission source , enabling scanning microscopes at high resolution. By the early 1980s improvements in mechanical stability as well as the use of higher accelerating voltages enabled imaging of materials at

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1166-422: The 1990s when it was discovered that mutations in the genes that code for lamins can be related to muscular dystrophies, cardiomyopathies, and neuropathies. Current research is being performed to develop treatment methods for the aforementioned laminopathies and to investigate the role lamins play in the aging process. The structure of lamins is composed of three units that are common among intermediate filaments:

1219-496: The A-type lamins. This suggests that the common ancestor of these lamin types was a B-type lamin. Due to their properties as a type of IF protein, lamins provide support for maintaining the shape of the nucleus. They also play an indirect role in anchoring the nucleus to the endoplasmic reticulum , forming a continuous unit within the cell. This is accomplished by lamin and lamin-interacting proteins (SUN1/SUN2) connecting with proteins on

1272-699: The LMNA gene, encoding Lamins A and C, can produce a series of disorders ranging from muscular dystrophies , neuropathies , cardiomyopathies , and premature ageing syndromes . Collectively, these conditions are known as laminopathies . One specific laminopathy is Hutchinson-Gilford progeria syndrome (HGPS), characterized by premature ageing. Those affected by the condition appear normal at birth, but show signs of premature ageing including hair-loss, thinness, joint abnormalities, and weak motor skills as they develop. Furthermore, health problems usually seen in older persons such as atherosclerosis and high blood pressure occur at

1325-399: The atomic scale. In the 1980s, the field emission gun became common for electron microscopes, improving the image quality due to the additional coherence and lower chromatic aberrations. The 2000s were marked by advancements in aberration-corrected electron microscopy, allowing for significant improvements in resolution and clarity of images. The original form of the electron microscope,

1378-660: The composition of the tail domain varies based on the type of lamin. However, all C-terminal domains contain a nuclear localization sequence (NLS). Similar to other IF proteins, lamins self-assemble into more complex structures. The basic unit of these structures is a coiled-coil dimer. The dimers arrange themselves in a head-to-tail manner, allowing for the formation of a protofilament. As these protofilaments aggregate, they form lamin filaments. Lamins of higher level organisms, such as vertebrates, continue to assemble into paracrystalline arrays. These complex structures allow nuclear lamins to perform their specialized functions in maintaining

1431-578: The data from the two modalities. This is commonly used to provide higher resolution contextual EM information about a fluorescently labelled structure. This correlative light and electron microscopy ( CLEM ) is one of a range of correlative workflows now available. Another example is high resolution mass spectrometry (ion microscopy), which has been used to provide correlative information about subcellular antibiotic localisation, data that would be difficult to obtain by other means. The initial role of electron microscopes in imaging two-dimensional slices (TEM) or

1484-458: The electron beam interacts with the specimen, it loses energy by a variety of mechanisms. These interactions lead to, among other events, emission of low-energy secondary electrons and high-energy backscattered electrons, light emission ( cathodoluminescence ) or X-ray emission, all of which provide signals carrying information about the properties of the specimen surface, such as its topography and composition. The image displayed by SEM represents

1537-505: The electrons hit the specimen in the STEM, but afterward in the TEM. The STEMs use of SEM-like beam rastering simplifies annular dark-field imaging , and other analytical techniques, but also means that image data is acquired in serial rather than in parallel fashion. The SEM produces images by probing the specimen with a focused electron beam that is scanned across the specimen ( raster scanning ). When

1590-612: The final fifteen residues by a zinc metalloprotease. The very first modification involving farnesylation of prelamin A is crucial to the development of mature lamin A. Isoform lamin C does not undergo posttranslational modifications. Some studies have demonstrated that lamins A and C are not required for the formation of the nuclear lamina, yet disruptions in the LMNA gene can contribute to physical and mental limitations. B-type lamins are characterized by an acidic isoelectric point, and they are typically expressed in every cell. As with A-type lamins, there are multiple isoforms of B-type lamins,

1643-413: The first electron microscope. (Max Knoll died in 1969, so did not receive a share of the 1986 Nobel prize for the invention of electron microscopes.) Apparently independent of this effort was work at Siemens-Schuckert by Reinhold Rüdenberg . According to patent law (U.S. Patent No. 2058914 and 2070318, both filed in 1932), he is the inventor of the electron microscope, but it is not clear when he had

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1696-457: The inner nuclear membrane. This disrupts the mechanical stability of the nucleus, resulting in a higher rate of cell death and therefore a higher rate of aging. Current studies are investigating the effects of farnesyl-transferase inhibitors (FTIs) to see if farnesyl attachment can be inhibited during posttranslational modification of prelamin A in order to treat patients with HGPS. Some laminopathies affect heart muscle . These mutations cause

1749-728: The molecules that make up air would scatter the electrons. An exception is liquid-phase electron microscopy using either a closed liquid cell or an environmental chamber, for example, in the environmental scanning electron microscope , which allows hydrated samples to be viewed in a low-pressure (up to 20  Torr or 2.7 kPa) wet environment. Various techniques for in situ electron microscopy of gaseous samples have been developed. Scanning electron microscopes operating in conventional high-vacuum mode usually image conductive specimens; therefore non-conductive materials require conductive coating (gold/palladium alloy, carbon, osmium, etc.). The low-voltage mode of modern microscopes makes possible

1802-635: The most common being lamin B1 and lamin B2 . They are produced from two separate genes, LMNB1 and LMNB2 . Similar to prelamin A, B-type lamins also contain a CaaX motif at the carboxyl-terminus. This marker triggers the same sequence of posttranslational modifications previously described for prelamin A except for the final cleavage step involving a zinc metalloprotease. Further investigations of B-type lamins across multiple species have found evidence that supports that B-type lamins existed before A-type lamins. This stems from

1855-508: The nuclear envelope under the inner nuclear membrane. It wasn't until 1986 that an analysis of lamin cDNA clones across a variety of species supported that lamins belong to the intermediate filament (IF) protein family. Further investigations found evidence that supports that all IF proteins arose from a common lamin-like ancestor. This theory is based on the observation that organisms that contain IF proteins necessarily contain lamins as well; however,

1908-569: The nuclear envelope. This allows chromatin to condense and the DNA to be replicated. After chromosome segregation, dephosphorylation of nuclear lamins by a phosphatase promotes reassembly of the nuclear envelope. Apoptosis is a highly organized process of programmed cell death. Lamins are crucial targets for this process due to their close associations with chromatin and the nuclear envelope. Apoptotic enzymes called caspases target lamins and cleave both A- and B-types. This allows chromatin to separate from

1961-443: The nuclear lamina in order to be condensed. As apoptosis continues, cell structures slowly shrink into compartmentalized "blebs." Finally, these apoptotic bodies are digested by phagocytes . Studies of apoptosis involving mutant A- and B-type lamins that are resistant to cleavage by caspases show decreased DNA condensation and apoptotic “blebbing” formation, thereby underscoring the important role of lamins in apoptosis. Mutations in

2014-714: The observation of non-conductive specimens without coating. Non-conductive materials can be imaged also by a variable pressure (or environmental) scanning electron microscope. Small, stable specimens such as carbon nanotubes , diatom frustules and small mineral crystals (asbestos fibres, for example) require no special treatment before being examined in the electron microscope. Samples of hydrated materials, including almost all biological specimens, have to be prepared in various ways to stabilize them, reduce their thickness (ultrathin sectioning) and increase their electron optical contrast (staining). These processes may result in artifacts , but these can usually be identified by comparing

2067-453: The other isoform. A CaaX motif is found within the unique residues in prelamin A. Due to the presence of the CaaX motif, prelamin A undergoes a series of posttranslational modifications to become mature lamin A. These steps include farnesylation of the carboxyl-terminal cysteine, endoproteolytic release of the terminal amino acids, carboxymethalation of the accessible farnesylcysteine, and removal of

2120-410: The outer nuclear membrane. These proteins in turn interact with cytoskeletal elements of the endoplasmic reticulum , forming a strong complex that can withstand mechanical stress. Nuclei that lack lamins or have mutated versions have a deformed shape and do not function properly. During mitosis, lamins are phosphorylated by Mitosis-Promoting Factor (MPF), which drives the disassembly of the lamina and

2173-678: The output is essentially a sequence of images through a specimen block that can be digitally aligned in sequence and thus reconstructed into a volume EM dataset. The increased volume available in these methods has expanded the capability of electron microscopy to address new questions, such as mapping neural connectivity in the brain, and membrane contact sites between organelles. Electron microscopes are expensive to build and maintain. Microscopes designed to achieve high resolutions must be housed in stable buildings (sometimes underground) with special services such as magnetic field canceling systems. The samples largely have to be viewed in vacuum , as

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2226-559: The physicist Leó Szilárd tried in 1928 to convince him to build an electron microscope, for which Szilárd had filed a patent. To this day the issue of who invented the transmission electron microscope is controversial. In 1928, at the Technische Hochschule in Charlottenburg (now Technische Universität Berlin ), Adolf Matthias (Professor of High Voltage Technology and Electrical Installations) appointed Max Knoll to lead

2279-480: The presence of lamins is not a requirement for simultaneously containing IF proteins. Furthermore, sequence comparisons between lamins and IF proteins support that an amino-acid sequence that is characteristic of lamins is found in early forms of IF proteins. This sequence is lost in later forms of IF proteins, suggesting that the structure of later intermediate filaments diverged. After this research, investigations of lamins slowed. Studies of lamins became more popular in

2332-473: The resolution in high-resolution transmission electron microscopy (HRTEM) to below 0.5 angstrom (50 picometres ), enabling magnifications above 50 million times. The ability of HRTEM to determine the positions of atoms within materials is useful for nano-technologies research and development. The STEM rasters a focused incident probe across a specimen. The high resolution of the TEM is thus possible in STEM. The focusing action (and aberrations) occur before

2385-489: The resolution of an optical (light) microscope. Four years later, in 1937, Siemens financed the work of Ernst Ruska and Bodo von Borries , and employed Helmut Ruska , Ernst's brother, to develop applications for the microscope, especially with biological specimens. Also in 1937, Manfred von Ardenne pioneered the scanning electron microscope . Siemens produced the first commercial electron microscope in 1938. The first North American electron microscopes were constructed in

2438-495: The results obtained by using radically different specimen preparation methods. Since the 1980s, analysis of cryofixed , vitrified specimens has also become increasingly used by scientists, further confirming the validity of this technique. Lamin B1 2KPW , 3JT0 , 3TYY , 3UMN 4001 16906 ENSG00000113368 ENSMUSG00000024590 P20700 P14733 NM_001198557 NM_005573 NM_010721 NP_001185486 NP_005564 NP_034851 Lamin-B1

2491-474: The shape of the nucleus as well as roles during mitosis and apoptosis. Lamins are divided into two major categories: A- and B-types. These subdivisions are based on similarities in cDNA sequences, structural features, isoelectric points, and expression trends. A-type lamins are characterized by a neutral isoelectric point , and they are typically displayed during later stages of embryonic development. Expressed in differentiated cells, A-type lamins originate from

2544-454: The signal in SEM, non-conductive samples (e.g. biological samples as in figure) can be sputter-coated in a thin film of metal. Materials to be viewed in a transmission electron microscope may require processing to produce a suitable sample. The technique required varies depending on the specimen and the analysis required: In their most common configurations, electron microscopes produce images with

2597-448: The similarity in structure of B-type lamins between invertebrates and vertebrates. Furthermore, organisms that only contain a single lamin contain a B-type lamin. Other studies that have investigated the structural similarities and differences between A- and B-type lamins have found that the positions of introns/exons in B-type lamins have been conserved in A-type lamins, with more variations in

2650-439: The structure of the specimen that is magnified by lenses of the microscope. The spatial variation in this information (the "image") may be viewed by projecting the magnified electron image onto a detector . For example, the image may be viewed directly by an operator using a fluorescent viewing screen coated with a phosphor or scintillator material such as zinc sulfide . A high-resolution phosphor may also be coupled by means of

2703-502: The two B type proteins, B1. Lamin B, along with heterochromatin , is anchored to the inner surface of the nuclear membrane by the lamin B receptor . LMNB1 has been shown to interact with Thymopoietin . When double-strand breaks are induced in DNA by ionizing radiation , lamin B1 promotes repair of the breaks , as well as cell survival, by maintaining the level of the RAD51 protein that

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2756-564: The varying intensity of any of these signals into the image in a position corresponding to the position of the beam on the specimen when the signal was generated. SEMs are different from TEMs in that they use electrons with much lower energy, generally below 20 keV, while TEMs generally use electrons with energies in the range of 80-300 keV. Thus, the electron sources and optics of the two microscopes have different designs, and they are normally separate instruments. Transmission electron microscopes can be used in electron diffraction mode where

2809-443: The z axis), a series of images taken through the sample depth can be used. For example, ribbons of serial sections can be imaged in a TEM as described above, and when thicker sections are used, serial TEM tomography can be used to increase the z-resolution. More recently, back scattered electron (BSE) images can be acquired of a larger series of sections collected on silicon wafers, known as SEM array tomography. An alternative approach

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