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Horseradish peroxidase is also commonly used in techniques such as ELISA and Immunohistochemistry due to its monomeric nature and the ease with which it produces coloured products. Peroxidase, a heme-containing oxidoreductase, is a commercially important enzyme which catalyses the reductive cleavage of hydrogen peroxide by an electron donor.
TMB can act as a hydrogen donor for the reduction of hydrogen peroxide to water by peroxidase enzymes such as horseradish peroxidase.. Shows the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB) to 3,3',5,5'-tetramethylbenzidine diimine
[28] [29] ELISA can also be used in toxicology as a rapid presumptive screen for certain classes of drugs. Enzyme-linked immunosorbent assay plate. The ELISA was the first screening test widely used for HIV because of its high sensitivity. In an ELISA, a person's serum is diluted 400 times and applied to a plate to which HIV antigens are attached.
As previously described, enzymes such as horseradish peroxidase or alkaline phosphatase are commonly used to catalyse reactions that give a coloured or chemiluminescent product. Fluorescent molecules can be visualised using fluorescence microscopy or confocal microscopy. [citation needed]
ELISA plate showing various cortisol levels. Enzymes used in ELISAs include horseradish peroxidase (HRP), alkaline phosphatase (AP) or glucose oxidase. These enzymes allow for detection often because they produce an observable color change in the presence of certain reagents.
Horseradish peroxidase (HRP) is a common enzyme utilized in ELISA schemes due to its ability to amplify signal and increase assay sensitivity. There are many variations, or types of ELISA assays but they can generally be classified as either indirect, competitive, sandwich or reverse. [21]