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Assay Method Information

Assay Name:  FRET-Based Z'-Lyte Assay
Description:  ABL activity assays were performed in 384-well plate using the FRET-based Z'-Lyteassay system and Tyr-2 peptide substrate according to the manufacturer's instructions (Invitrogen). Briefly, 10 μL reaction mixtures per well contain 12 μM ATP, 2 μM Tyr2 peptide substrate in 50 mm HEPES pH 7.5, 0.01% BRIJ-35, 10 mm MgCl2, 1 mm EGTA, 0.0247 μg/mL ABL1 and inhibitors as appropriate. The reaction was performed at room temperature for 2 hours, and 5 μL development reagent was then added for further 2-hour room temperature incubation followed by the addition of 5 μL of stop solution. Fluorescence signal ratio of 445 nm (coumarin) to 520 nm (fluorescin) was examined on EnVision Multilabel Reader (Perkin Elmer, Inc.).
Affinity data for this assay
 

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Last update November 1, 2007
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