Cell Unroofing
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Cell Unroofing
Cell unroofing is any of various methods to isolate and expose the cell membrane of cells. Differently from the more common membrane extraction protocols performed with multiple steps of centrifugation (which goal is to separate the membrane fraction from a cell lysate), in ''cell unroofing'' the aim is to tear and preserve patches of the plasma membrane in order to perform ''in situ'' experiments using (microscopy and biomedical spectroscopy). History The first observation the bi-layer cell membrane was made in 1959 on a section of a cell using the electron microscope. But the first micrograph of the internal side of a cell dates back to 1977 by M.V. Nermut. Professor John Heuser made substantial contributions in the field, imaging the detailed internal structure of the membrane and the cytoskeleton bound to it with extensive use of the electron microscope. It was only after the development of the atomic force microscope operated in liquid that it was possible to image the ce ...
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Sketch Cell Unroofing
Sketch or Sketches may refer to: * Sketch (drawing), a rapidly executed freehand drawing that is not usually intended as a finished work Arts, entertainment and media * Sketch comedy, a series of short scenes or vignettes called sketches Film and television * ''Sketch'' (2007 film), a Malayalam film * ''Sketch'' (2018 film), a Tamil film * ''Sketch'' (TV series), a 2018 South Korean series * "Sketch", a 2008 episode of ''Skins'' ** Sketch (''Skins'' character) * Sketch with Kevin McDonald, a 2006 CBC television special Literature * Sketch story, or sketch, a very short piece of writing * ''Daily Sketch'', a British newspaper 1909–1971 * ''The Sketch'', a British illustrated weekly journal 1893–1959 Music * Sketch (music), an informal document prepared by a composer to assist in composition * The Sketches, a Pakistani Sufi folk rock band * ''Sketch'' (album), by Ex Norwegian, 2011 * ''Sketch'' (EP), by Hyomin, 2016 * ''Sketches'' (album), by Bert Jansch, 1990 * ...
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Cell Membrane
The cell membrane (also known as the plasma membrane (PM) or cytoplasmic membrane, and historically referred to as the plasmalemma) is a biological membrane that separates and protects the interior of all cells from the outside environment (the extracellular space). The cell membrane consists of a lipid bilayer, made up of two layers of phospholipids with cholesterols (a lipid component) interspersed between them, maintaining appropriate membrane fluidity at various temperatures. The membrane also contains membrane proteins, including integral proteins that span the membrane and serve as membrane transporters, and peripheral proteins that loosely attach to the outer (peripheral) side of the cell membrane, acting as enzymes to facilitate interaction with the cell's environment. Glycolipids embedded in the outer lipid layer serve a similar purpose. The cell membrane controls the movement of substances in and out of cells and organelles, being selectively permeable to ions a ...
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Cell (biology)
The cell is the basic structural and functional unit of life forms. Every cell consists of a cytoplasm enclosed within a membrane, and contains many biomolecules such as proteins, DNA and RNA, as well as many small molecules of nutrients and metabolites.Cell Movements and the Shaping of the Vertebrate Body
in Chapter 21 of
Molecular Biology of the Cell
'' fourth edition, edited by Bruce Alberts (2002) published by Garland Science. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos. It is also common to describe small molecules such as ...
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Lysis
Lysis ( ) is the breaking down of the membrane of a cell, often by viral, enzymic, or osmotic (that is, "lytic" ) mechanisms that compromise its integrity. A fluid containing the contents of lysed cells is called a ''lysate''. In molecular biology, biochemistry, and cell biology laboratories, cell cultures may be subjected to lysis in the process of purifying their components, as in protein purification, DNA extraction, RNA extraction, or in purifying organelles. Many species of bacteria are subject to lysis by the enzyme lysozyme, found in animal saliva, egg white, and other secretions. Phage lytic enzymes (lysins) produced during bacteriophage infection are responsible for the ability of these viruses to lyse bacterial cells. Penicillin and related β-lactam antibiotics cause the death of bacteria through enzyme-mediated lysis that occurs after the drug causes the bacterium to form a defective cell wall. If the cell wall is completely lost and the penicillin was used ...
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Biomedical Spectroscopy
Biomedical spectroscopy is a multidisciplinary research field involving spectroscopic tools for applications in the field of biomedical science. Vibrational spectroscopy such as Raman or infrared spectroscopy is used to determine the chemical composition of a material based on detection of vibrational modes of constituent molecules. Some spectroscopic methods are routinely used in clinical settings for diagnosis of disease; an example is Magnetic resonance imaging Magnetic resonance imaging (MRI) is a medical imaging technique used in radiology to form pictures of the anatomy and the physiological processes of the body. MRI scanners use strong magnetic fields, magnetic field gradients, and radio wave ... (MRI). Fourier transform infrared (FTIR) spectroscopic imaging is a form of chemical imaging for which the contrast is provided by composition of the material. References {{reflist Spectroscopy ...
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John Heuser
John E. Heuser (born August 29, 1942) is an American Professor of Biophysics in the department of Cell Biology and Physiology at the Washington University School of Medicine as well as a Professor at the Institute for Integrated Cell-Material Sciences (iCeMS) at Kyoto University. Heuser created quick-freeze deep-etch electron microscopy (a variant of cell unroofing), a pioneering technique that lets biologists take detailed pictures of fleeting events inside living cells. For decades, Heuser has used this technique to capture details of the molecular mechanisms that underlie many basic biological activities, including nerve cell signal transmission, muscle contraction, and most recently, the fusion of viruses with cells during the spread of infection. He compares quick-freeze deep-etch electron microscopy to using a stroboscopic flash to freeze the action in a photograph. To make it possible to image the frozen sample with an electron microscope, Heuser adds an ultra-thin fil ...
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Cytoskeleton
The cytoskeleton is a complex, dynamic network of interlinking protein filaments present in the cytoplasm of all cells, including those of bacteria and archaea. In eukaryotes, it extends from the cell nucleus to the cell membrane and is composed of similar proteins in the various organisms. It is composed of three main components, microfilaments, intermediate filaments and microtubules, and these are all capable of rapid growth or disassembly dependent on the cell's requirements. A multitude of functions can be performed by the cytoskeleton. Its primary function is to give the cell its shape and mechanical resistance to deformation, and through association with extracellular connective tissue and other cells it stabilizes entire tissues. The cytoskeleton can also contract, thereby deforming the cell and the cell's environment and allowing cells to migrate. Moreover, it is involved in many cell signaling pathways and in the uptake of extracellular material ( endocytosis), the ...
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Atomic Force Microscopy
Atomic force microscopy (AFM) or scanning force microscopy (SFM) is a very-high-resolution type of scanning probe microscopy (SPM), with demonstrated resolution on the order of fractions of a nanometer, more than 1000 times better than the optical diffraction limit. Overview Atomic force microscopy (AFM) is a type of scanning probe microscopy (SPM), with demonstrated resolution on the order of fractions of a nanometer, more than 1000 times better than the optical diffraction limit. The information is gathered by "feeling" or "touching" the surface with a mechanical probe. Piezoelectric elements that facilitate tiny but accurate and precise movements on (electronic) command enable precise scanning. Despite the name, the Atomic Force Microscope does not use the Nuclear force. Abilities The AFM has three major abilities: force measurement, topographic imaging, and manipulation. In force measurement, AFMs can be used to measure the forces between the probe and the sample as ...
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Electron Microscope
An electron microscope is a microscope that uses a beam of accelerated electrons as a source of illumination. As the wavelength of an electron can be up to 100,000 times shorter than that of visible light photons, electron microscopes have a higher resolving power than light microscopes and can reveal the structure of smaller objects. A scanning transmission electron microscope has achieved better than 50  pm resolution in annular dark-field imaging mode and magnifications of up to about 10,000,000× whereas most light microscopes are limited by diffraction to about 200  nm resolution and useful magnifications below 2000×. Electron microscopes use shaped magnetic fields to form electron optical lens systems that are analogous to the glass lenses of an optical light microscope. Electron microscopes are used to investigate the ultrastructure of a wide range of biological and inorganic specimens including microorganisms, cells, large molecules, biopsy samples, ...
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Electron Microscopy
An electron microscope is a microscope that uses a beam of accelerated electrons as a source of illumination. As the wavelength of an electron can be up to 100,000 times shorter than that of visible light photons, electron microscopes have a higher resolving power than light microscopes and can reveal the structure of smaller objects. A scanning transmission electron microscope has achieved better than 50  pm resolution in annular dark-field imaging mode and magnifications of up to about 10,000,000× whereas most light microscopes are limited by diffraction to about 200  nm resolution and useful magnifications below 2000×. Electron microscopes use shaped magnetic fields to form electron optical lens systems that are analogous to the glass lenses of an optical light microscope. Electron microscopes are used to investigate the ultrastructure of a wide range of biological and inorganic specimens including microorganisms, cells, large molecules, biopsy samples, ...
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Cryofixation
Cryofixation is a technique for fixation or stabilisation of biological materials as the first step in specimen preparation for electron microscopy and cryo-electron microscopy. Typical specimens for cryofixation include small samples of plant or animal tissue, cell suspensions of microorganisms or cultured cells, suspensions of viruses or virus capsids and samples of purified macromolecules, especially proteins. Cryo fixation Types 1.Freezing-drying 2.Freezing-substitution 3.freezing-etching Plunge freezing The method involves ultra-rapid cooling of small tissue or cell samples to the temperature of liquid nitrogen (−196 °C) or below, stopping all motion and metabolic activity and preserving the internal structure by freezing all fluid phases solid. Typically, a sample is plunged into liquid nitrogen or into liquid ethane or liquid propane in a container cooled by liquid nitrogen. The ultimate objective is to freeze the specimen so rapidly (at 104 to 106 K per secon ...
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Sonication
A sonicator at the Weizmann Institute of Science during sonicationSonication is the act of applying sound energy to agitate particles in a sample, for various purposes such as the extraction of multiple compounds from plants, microalgae and seaweeds. Ultrasonic frequencies (> 20 kHz) are usually used, leading to the process also being known as ultrasonication or ultra-sonication. In the laboratory, it is usually applied using an ''ultrasonic bath'' or an ''ultrasonic probe'', colloquially known as a ''sonicator''. In a paper machine, an ultrasonic foil can distribute cellulose fibres more uniformly and strengthen the paper. Effects Sonication has numerous effects, both chemical and physical. The chemical effects of ultrasound are concerned with understanding the effect of sonic waves on chemical systems, this is called sonochemistry. The chemical effects of ultrasound do not come from a direct interaction with molecular species. Studies have shown that no direct coupl ...
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