Western Blot Normalization
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Western Blot Normalization
Normalization of Western blot data is an analytical step that is performed to compare the relative abundance of a specific protein across the lanes of a blot or gel under diverse experimental treatments, or across tissues or developmental stages. The overall goal of normalization is to minimize effects arising from variations in experimental errors, such as inconsistent sample preparation, unequal sample loading across gel lanes, or uneven protein transfer, which can compromise the conclusions that can be obtained from Western blot data. Currently, there are two methods for normalizing Western blot data: (i) housekeeping protein normalization and (ii) total protein normalization. Procedure Normalization occurs directly on either the gel or the blotting membrane. First, the stained gel or blot is imaged, a rectangle is drawn around the target protein in each lane, and the signal intensity inside the rectangle is measured. The signal intensity obtained can then be normalized with ...
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Western Blot
The western blot (sometimes called the protein immunoblot), or western blotting, is a widely used analytical technique in molecular biology and immunogenetics to detect specific proteins in a sample of tissue homogenate or extract. Besides detecting the proteins, this technique is also utilized to visualize, distinguish, and quantify the different proteins in a complicated protein combination. Western blot technique uses three elements to achieve its task of separating a specific protein from a complex: separation by size, transfer of protein to a solid support, and marking target protein using a primary and secondary antibody to visualize. A synthetic or animal-derived antibody (known as the primary antibody) is created that recognizes and binds to a specific target protein. The electrophoresis membrane is washed in a solution containing the primary antibody, before excess antibody is washed off. A secondary antibody is added which recognizes and binds to the primary antibody ...
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Molecular Mass
The molecular mass (''m'') is the mass of a given molecule: it is measured in daltons (Da or u). Different molecules of the same compound may have different molecular masses because they contain different isotopes of an element. The related quantity relative molecular mass, as defined by IUPAC, is the ratio of the mass of a molecule to the unified atomic mass unit (also known as the dalton) and is unitless. The molecular mass and relative molecular mass are distinct from but related to the molar mass. The molar mass is defined as the mass of a given substance divided by the amount of a substance and is expressed in g/mol. That makes the molar mass an average of many particles or molecules, and the molecular mass the mass of one specific particle or molecule. The molar mass is usually the more appropriate figure when dealing with macroscopic (weigh-able) quantities of a substance. The definition of molecular weight is most authoritatively synonymous with relative molecular mass; ho ...
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Amido Black 10B
Amido black 10B is an amino acid staining azo dye used in biochemical research to stain for total protein on transferred membrane blots, such as the western blot. It is also used in criminal investigations to detect blood present with latent fingerprints. It stains the proteins in blood a blue-black color. Amido Black can be either methanol or water based as it readily dissolves in both. With picric acid, in a van Gieson procedure, it can be used to stain collagen and reticulin. See also *Western blot normalization Normalization of Western blot data is an analytical step that is performed to compare the relative abundance of a specific protein across the lanes of a blot or gel under diverse experimental treatments, or across tissues or developmental stages. T ... References External linksMSDS at Oxford University {{DEFAULTSORT:Amido Black 10b Azo dyes Naphthalenesulfonates Organic sodium salts 1-Naphthols Nitrobenzenes Aromatic amines Acid dyes ...
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Camera
A camera is an Optics, optical instrument that can capture an image. Most cameras can capture 2D images, with some more advanced models being able to capture 3D images. At a basic level, most cameras consist of sealed boxes (the camera body), with a small hole (the aperture) that allows light to pass through in order to capture an image on a light-sensitive surface (usually a Image sensor, digital sensor or photographic film). Cameras have various mechanisms to control how the light falls onto the light-sensitive surface. Lenses focus the light entering the camera, and the aperture can be narrowed or widened. A Shutter (photography), shutter mechanism determines the amount of time the photosensitive surface is exposed to the light. The still image camera is the main instrument in the art of photography. Captured images may be reproduced later as part of the process of photography, digital imaging, or photographic printing. Similar artistic fields in the moving-image camera dom ...
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Charge-coupled Device
A charge-coupled device (CCD) is an integrated circuit containing an array of linked, or coupled, capacitors. Under the control of an external circuit, each capacitor can transfer its electric charge to a neighboring capacitor. CCD sensors are a major technology used in digital imaging. In a CCD image sensor, pixels are represented by p-doped metal–oxide–semiconductor (MOS) capacitors. These MOS capacitors, the basic building blocks of a CCD, are biased above the threshold for inversion when image acquisition begins, allowing the conversion of incoming photons into electron charges at the semiconductor-oxide interface; the CCD is then used to read out these charges. Although CCDs are not the only technology to allow for light detection, CCD image sensors are widely used in professional, medical, and scientific applications where high-quality image data are required. In applications with less exacting quality demands, such as consumer and professional digital cameras, act ...
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Coomassie Brilliant Blue
Coomassie brilliant blue is the name of two similar triphenylmethane dyes that were developed for use in the textile industry but are now commonly used for staining proteins in analytical biochemistry. Coomassie brilliant blue G-250 differs from Coomassie brilliant blue R-250 by the addition of two methyl groups. The name "Coomassie" is a registered trademark of Imperial Chemical Industries. Name and discovery The name Coomassie was adopted at the end of the 19th century as a trade name by the Blackley-based dye manufacturer Levinstein Ltd, in marketing a range of acid wool dyes. In 1896 during the Fourth Anglo–Ashanti War, British forces had occupied the town of Coomassie (modern-day Kumasi in Ghana). In 1918 Levinstein Ltd became part of British Dyestuffs, which in 1926 became part of Imperial Chemical Industries. Although ICI still owns the Coomassie trademark, the company no longer manufactures the dyes. The blue disulfonated triphenylmethane dyes were first produced in ...
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Exogeny
In a variety of contexts, exogeny or exogeneity () is the fact of an action or object originating externally. It contrasts with endogeneity or endogeny, the fact of being influenced within a system. Economics In an economic model, an exogenous change is one that comes from outside the model and is unexplained by the model. Such changes of an economic model from outside factors can include the influence of technology, in which this had previously been noted as an exogenous factor, but has rather been noted as a factor that can depict economic forces as a whole. In economic sociology, Project IDEA (Interdisciplinary Dimensions of Economic Analysis) gave notion to understanding the exogenous factors that play a role within economic theory. Developed from the International Social Science Council (ISSC) in the year of 1982, Project IDEA was founded to gather ideas from economists and sociologists in order to conceptualize what economic sociology incorporates, as they have sought ...
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Western Blot
The western blot (sometimes called the protein immunoblot), or western blotting, is a widely used analytical technique in molecular biology and immunogenetics to detect specific proteins in a sample of tissue homogenate or extract. Besides detecting the proteins, this technique is also utilized to visualize, distinguish, and quantify the different proteins in a complicated protein combination. Western blot technique uses three elements to achieve its task of separating a specific protein from a complex: separation by size, transfer of protein to a solid support, and marking target protein using a primary and secondary antibody to visualize. A synthetic or animal-derived antibody (known as the primary antibody) is created that recognizes and binds to a specific target protein. The electrophoresis membrane is washed in a solution containing the primary antibody, before excess antibody is washed off. A secondary antibody is added which recognizes and binds to the primary antibody ...
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Protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, responding to stimuli, providing structure to cells and organisms, and transporting molecules from one location to another. Proteins differ from one another primarily in their sequence of amino acids, which is dictated by the nucleotide sequence of their genes, and which usually results in protein folding into a specific 3D structure that determines its activity. A linear chain of amino acid residues is called a polypeptide. A protein contains at least one long polypeptide. Short polypeptides, containing less than 20–30 residues, are rarely considered to be proteins and are commonly called peptides. The individual amino acid residues are bonded together by peptide bonds and adjacent amino acid residues. The sequence of amino acid residue ...
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RPLP1
60S acidic ribosomal protein P1 is a protein that in humans is encoded by the ''RPLP1'' gene. Function Ribosomes, the organelles that catalyze protein synthesis, consist of a small 40S subunit and a large 60S subunit. Together these subunits are composed of 4 RNA species and approximately 80 structurally distinct proteins. This gene encodes a ribosomal phosphoprotein that is a component of the 60S subunit. The protein, which is a functional equivalent of the Escherichia coli L7/ L12 ribosomal protein, belongs to the L12P family of ribosomal proteins. It plays an important role in the elongation step of protein synthesis. Unlike most ribosomal proteins, which are basic, the encoded protein is acidic. Its C-terminal end is nearly identical to the C-terminal ends of the ribosomal phosphoproteins P0 and P2. The P1 protein can interact with P0 and P2 to form a pentameric complex consisting of P1 and P2 dimers, and a P0 monomer. The protein is located in the cytoplasm. Two alterna ...
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Hypoxanthine-guanine Phosphoribosyltransferase
Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is an enzyme encoded in humans by the ''HPRT1'' gene. HGPRT is a transferase that catalyzes conversion of hypoxanthine to inosine monophosphate and guanine to guanosine monophosphate. This reaction transfers the 5-phosphoribosyl group from 5-phosphoribosyl 1-pyrophosphate (PRPP) to the purine. HGPRT plays a central role in the generation of purine nucleotides through the purine salvage pathway. Function HGPRT catalyzes the following reactions: HGPRTase functions primarily to salvage purines from degraded DNA to reintroduce into purine synthetic pathways. In this role, it catalyzes the reaction between guanine and phosphoribosyl pyrophosphate (PRPP) to form GMP, or between hypoxanthine and phosphoribosyl pyrophosphate (PRPP) to form inosine monophosphate. Substrates and inhibitors Comparative homology modelling of this enzyme in '' L. donovani'' suggest that among all of the computationally screened compounds, p ...
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