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Assay Method Information

Assay Name:  In Vitor Src Tyrosine Kinase Activity Assay
Description:  Kinase activity (Src-family kinases, Hck, Lyn, Fyn, and c-Src) and the effect of the molecules were determined by ProFluor Src-Family Kinase Assay Kit (Promega). A titration assay was performed for each kinase to determine the amount of the enzyme that results in approximately 80% phosphorylation as suggested by the manufacturer. The compounds were dissolved in 10% DMSO and tested at 1, 10 and 100 μM concentrations. Briefly, the molecules were mixed with a reaction buffer that included a specific substrate for Src-family kinases (R110), a control substrate and the kinase. The reaction was initiated by the addition of ATP. After incubating the 96-well reaction plate at 22C for 1 h, protease solution was added to each well and the plate was incubated for 1 h. The fluorescence of the liberated R110 was read at an excitation wavelength of 485 nm and emission wavelength of 530 nm.
Affinity data for this assay
 

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Last update November 1, 2007
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