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The enzyme alkaline phosphatase (ALP, alkaline phenyl phosphatase, also abbreviated PhoA) is a phosphatase with the physiological role of dephosphorylating compounds. The enzyme is found across a multitude of organisms, prokaryotes and eukaryotes alike, with the same general function, but in different structural forms suitable to the environment they function in. Alkaline phosphatase is found ...
5-Bromo-4-chloro-3-indolyl phosphate (BCIP, X-phosphate, XP) is an artificial chromogenic substrate used for the sensitive colorimetric detection of alkaline phosphatase activity. It is, for example, used in immunoblotting , in situ hybridization , and immunohistochemistry , often in combination with nitro blue tetrazolium chloride (NBT).
Specific activity gives a measurement of enzyme purity in the mixture. It is the micro moles of product formed by an enzyme in a given amount of time (minutes) under given conditions per milligram of total proteins. Specific activity is equal to the rate of reaction multiplied by the volume of reaction divided by the mass of total protein.
The relative activity of a species i, denoted a i, is defined [4] [5] as: = where μ i is the (molar) chemical potential of the species i under the conditions of interest, μ o i is the (molar) chemical potential of that species under some defined set of standard conditions, R is the gas constant, T is the thermodynamic temperature and e is the exponential constant.
One application is in activity stains in gel electrophoresis, such as with the mitochondrial electron transport chain complexes. [ 2 ] Nitro blue tetrazolium is used in a diagnostic test, [ 3 ] particularly for chronic granulomatous disease and other diseases of phagocyte function.
ALP activity is significantly increased in the third trimester of pregnancy. [11] This is due to increased synthesis from the placenta as well as increased synthesis in the liver induced by large amounts of estrogens. [11] [12] [13] Levels in the third trimester can be as much as 2-fold greater than in non-pregnant women. [11]
However, when experiments have been carried out in-vivo, phosphatase enzymes have been shown to be incredibly specific. [3] In some cases, a protein phosphatase (i.e. one defined by its recognition of protein substrates) can catalyze the dephosphorylation of nonprotein substrates. [ 4 ]
In the initially published VCC experiments [1] only the internal 60 wells of the plate were used, since evaporation changed the volume of the edge wells during the 12-hour incubation. However, all 96 wells of the 96-well plate can be used for the experiment as long as the edge of the plate is wrapped with a piece of Parafilm M six squares long ...