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Assay Method Information

Assay Name:  uHTS identification of compounds inhibiting the binding between the RUNX1 Runt domain and CBFb-SMMHC via a fluorescence resonance energy transfer (FRET) assay.
Description:  This assay is to use HTS to identify inhibitors of the protein-protein interaction between the RUNX1 Runt domain and CBFbeta-SMMHC, a potential therapeutic approach for inv(16) related leukemia. This is accomplished by using a fluorescence resonance energy transfer (FRET) assay. When added together Cerulean-Runt domain and Venus-CBFbeta-SMMHC will bind and allow for energy transfer. If an inhibitor is present, binding will not occur and the FRET signal will be abated. The fluorescent CBF alpha/beta complex is incubated in black 1536-well plate at room temperature for 1h. Positive control (PC) solution contained untagged CBFbeta-SMMHC. After incubation, the plate is read on a BMG Pherastar 422ex, 530em and 480em. The FRET signal is determined as the ratio of 530/480 signals. IC50 value was determined using sigmoidal dose response equation.
Affinity data for this assay
 

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Last update November 1, 2007
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