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Streptamer
The Streptamer technology allows the reversible isolation and staining of antigen-specific T cells. This technology combines a current T cell isolation method with the Strep-tag technology. In principle, the T cells are separated by establishing a specific interaction between the T cell of interest and a molecule that is conjugated to a marker, which enables the isolation. The reversibility of this interaction and the low temperatures at which it is performed allows for the isolation and characterization of functional T cells. Because T cells remain phenotypically and functionally indistinguishable from untreated cells, this method offers modern strategies in clinical and basic T cell research. Classic methods in T cell research T cells play an important role in the adaptive immune system. They are capable of orchestrating, regulating and coordinating complex immune responses. A wide array of clinically relevant aspects are associated with the function or malfunction of T-cells: A ...
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Major Histocompatibility Complex
The major histocompatibility complex (MHC) is a large locus on vertebrate DNA containing a set of closely linked polymorphic genes that code for cell surface proteins essential for the adaptive immune system. These cell surface proteins are called MHC molecules. This locus got its name because it was discovered via the study of transplanted tissue compatibility. Later studies revealed that tissue rejection due to incompatibility is only a facet of the full function of MHC molecules: binding an antigen derived from self-proteins, or from pathogens, and bringing the antigen presentation to the cell surface for recognition by the appropriate T-cells. MHC molecules mediate the interactions of leukocytes, also called white blood cells (WBCs), with other leukocytes or with body cells. The MHC determines donor compatibility for organ transplant, as well as one's susceptibility to autoimmune diseases. In a cell, protein molecules of the host's own phenotype or of other biologic entities ...
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Strep-tag
The Strep-tag system is a method which allows the purification and detection of proteins by affinity chromatography. The Strep-tag II is a synthetic peptide consisting of eight amino acids (Tryptophan, Trp-Serine, Ser-Histidine, His-Proline, Pro-Gln-Phenylalanine, Phe-Glutamic acid, Glu-Lysine, Lys). This peptide sequence exhibits intrinsic affinity towards Strep-Tactin, a specifically engineered streptavidin, and can be N- or C- terminally fused to recombinant proteins. By exploiting the highly specific interaction, ''Strep''-tagged proteins can be isolated in one step from crude cell lysates. Because the ''Strep''-tag elutes under gentle, physiological conditions, it is especially suited for generation of functional proteins. Development and biochemistry of the Strep-tag Streptavidin is a tetrameric protein expressed in ''Streptomyces avidinii''. Because of Streptavidin's high affinity for vitamin H (biotin), Streptavidin is commonly used in the fields of molecular biology a ...
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Molecular Biology Techniques
A molecule is a group of two or more atoms held together by attractive forces known as chemical bonds; depending on context, the term may or may not include ions which satisfy this criterion. In quantum physics, organic chemistry, and biochemistry, the distinction from ions is dropped and ''molecule'' is often used when referring to polyatomic ions. A molecule may be homonuclear, that is, it consists of atoms of one chemical element, e.g. two atoms in the oxygen molecule (O2); or it may be heteronuclear, a chemical compound composed of more than one element, e.g. water (two hydrogen atoms and one oxygen atom; H2O). In the kinetic theory of gases, the term ''molecule'' is often used for any gaseous particle regardless of its composition. This relaxes the requirement that a molecule contains two or more atoms, since the noble gases are individual atoms. Atoms and complexes connected by non-covalent interactions, such as hydrogen bonds or ionic bonds, are typically not considere ...
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Biotechnology
Biotechnology is the integration of natural sciences and engineering sciences in order to achieve the application of organisms, cells, parts thereof and molecular analogues for products and services. The term ''biotechnology'' was first used by Károly Ereky in 1919, meaning the production of products from raw materials with the aid of living organisms. Definition The concept of biotechnology encompasses a wide range of procedures for modifying living organisms according to human purposes, going back to domestication of animals, cultivation of the plants, and "improvements" to these through breeding programs that employ artificial selection and hybridization. Modern usage also includes genetic engineering as well as cell and tissue culture technologies. The American Chemical Society defines biotechnology as the application of biological organisms, systems, or processes by various industries to learning about the science of life and the improvement of the value of materials ...
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Protein Methods
Protein methods are the techniques used to study proteins. There are experimental methods for studying proteins (e.g., for detecting proteins, for isolating and purifying proteins, and for characterizing the structure and function of proteins, often requiring that the protein first be purified). Computational methods typically use computer programs to analyze proteins. However, many experimental methods (e.g., mass spectrometry) require computational analysis of the raw data. Genetic methods Experimental analysis of proteins typically requires expression and purification of proteins. Expression is achieved by manipulating DNA that encodes the protein(s) of interest. Hence, protein analysis usually requires DNA methods, especially cloning. Some examples of genetic methods include conceptual translation, Site-directed mutagenesis, using a fusion protein, and matching allele with disease states. Some proteins have never been directly sequenced, however by translating codons from know ...
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Biotin
Biotin (or vitamin B7) is one of the B vitamins. It is involved in a wide range of metabolic processes, both in humans and in other organisms, primarily related to the utilization of fats, carbohydrates, and amino acids. The name ''biotin'', borrowed from the German , derives from the Ancient Greek word (; 'life') and the suffix "-in" (a suffix used in chemistry usually to indicate 'forming'). Chemical description Biotin is classified as a heterocyclic compound, with a sulfur-containing ring fused ureido and tetrahydrothiophene group. A C5-carboxylic acid side chain is appended to one of the rings. The ureido ring, containing the −N−CO−N− group, serves as the carbon dioxide carrier in carboxylation reactions. Biotin is a coenzyme for five carboxylase enzymes, which are involved in the catabolism of amino acids and fatty acids, synthesis of fatty acids, and gluconeogenesis. Biotinylation of histone proteins in nuclear chromatin plays a role in chromatin stability and g ...
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Flow Cytometry
Flow cytometry (FC) is a technique used to detect and measure physical and chemical characteristics of a population of cells or particles. In this process, a sample containing cells or particles is suspended in a fluid and injected into the flow cytometer instrument. The sample is focused to ideally flow one cell at a time through a laser beam, where the light scattered is characteristic to the cells and their components. Cells are often labeled with fluorescent markers so light is absorbed and then emitted in a band of wavelengths. Tens of thousands of cells can be quickly examined and the data gathered are processed by a computer. Flow cytometry is routinely used in basic research, clinical practice, and clinical trials. Uses for flow cytometry include: * Cell counting * Cell sorting * Determining cell characteristics and function * Detecting microorganisms * Biomarker detection * Protein engineering detection * Diagnosis of health disorders such as blood cancers * Measuring ...
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Fluorochromes
A fluorophore (or fluorochrome, similarly to a chromophore) is a fluorescence, fluorescent chemical compound that can re-emit light upon light excitation. Fluorophores typically contain several combined aromaticity, aromatic groups, or planar or cyclic molecules with several pi bond, π bonds. Fluorophores are sometimes used alone, as a dye tracing, tracer in fluids, as a dye for staining of certain structures, as a substrate of enzymes, or as a probe or indicator (when its fluorescence is affected by environmental aspects such as polarity or ions). More generally they are covalent bond, covalently bonded to a macromolecule, serving as a marker (or dye, or tag, or reporter) for affine or bioactive reagents (antibodies, peptides, nucleic acids). Fluorophores are notably used to stain tissues, cells, or materials in a variety of analytical methods, i.e., Fluorescence microscope, fluorescent imaging and fluorescence spectroscopy, spectroscopy. Fluorescein, via its amine-reactive i ...
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Avidity
In biochemistry, avidity refers to the accumulated strength of ''multiple'' affinities of individual non-covalent binding interactions, such as between a protein receptor and its ligand, and is commonly referred to as functional affinity. Avidity differs from affinity, which describes the strength of a ''single'' interaction. However, because individual binding events increase the likelihood of occurrence of other interactions (i.e., increase the local concentration of each binding partner in proximity to the binding site), avidity should not be thought of as the mere sum of its constituent affinities but as the combined effect of all affinities participating in the biomolecular interaction. A particular important aspect relates to the phenomenon of 'avidity entropy'. Biomolecules often form heterogenous complexes or homogeneous oligomers and multimers or polymers. If clustered proteins form an organized matrix, such as the clathrin-coat, the interaction is described as a matricity. ...
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Affinity (pharmacology)
In biochemistry and pharmacology, a ligand is a substance that forms a complex with a biomolecule to serve a biological purpose. The etymology stems from ''ligare'', which means 'to bind'. In protein-ligand binding, the ligand is usually a molecule which produces a signal by binding to a site on a target protein. The binding typically results in a change of conformational isomerism (conformation) of the target protein. In DNA-ligand binding studies, the ligand can be a small molecule, ion, or protein which binds to the DNA double helix. The relationship between ligand and binding partner is a function of charge, hydrophobicity, and molecular structure. Binding occurs by intermolecular forces, such as ionic bonds, hydrogen bonds and Van der Waals forces. The association or docking is actually reversible through dissociation. Measurably irreversible covalent bonding between a ligand and target molecule is atypical in biological systems. In contrast to the definition of liga ...
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T Cell Receptor
The T-cell receptor (TCR) is a protein complex found on the surface of T cells, or T lymphocytes, that is responsible for recognizing fragments of antigen as peptides bound to major histocompatibility complex (MHC) molecules. The binding between TCR and antigen peptides is of relatively low affinity and is degenerate: that is, many TCRs recognize the same antigen peptide and many antigen peptides are recognized by the same TCR. The TCR is composed of two different protein chains (that is, it is a heterodimer). In humans, in 95% of T cells the TCR consists of an alpha (α) chain and a beta (β) chain (encoded by '' TRA'' and ''TRB'', respectively), whereas in 5% of T cells the TCR consists of gamma and delta (γ/δ) chains (encoded by '' TRG'' and '' TRD'', respectively). This ratio changes during ontogeny and in diseased states (such as leukemia). It also differs between species. Orthologues of the 4 loci have been mapped in various species. Each locus can produce a vari ...
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