| Assay Method Information | |
| | FRET Assay |
| Description: | For the FRET measurements, the Cerulean-Runt (Runx1 41-190) and Venus-CBFbeta (1-141) fusion proteins were dialyzed into FRET assay buffer. Protein concentrations were determined by UV absorption at 433 nm for Cerulean-Runt and 513 nm for Venus-CBFbeta. Equal molar concentrations of the two proteins were mixed together, diluted to 150 nM with FRET assay buffer containing 0.1% BSA, and incubated for 1 hr. A PHERAstar microplate reader (BMG Labtech, Durham, NC, USA) was used to measure fluorescence (excitation at 433 nm and emission measured at 474 and 525 nm). For IC50 determinations, the ratios of the fluorescence intensities at 525 nm and 474 nm were plotted versus the log of compound concentration, and the resulting curve was fit to a sigmoidal curve using Origin 7.0 (MicroCal, Northampton, MA, USA). Mean values of IC50 originating from two independent measurements performed in duplicate together with the standard deviations are reported. |
| Affinity data for this assay | |
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