| Assay Method Information | |
| | Enzymologic Experiment of FGFR4 |
| Description: | In this experiment, the inhibitory effect of the compounds on FGFR4 kinase activity was tested by a fluorescence resonance energy transfer (TR-FRET) method, and the half maximal inhibitory concentration (IC50) of the compounds on the FGFR4 kinase activity was determined. 1) 1 5 μL of FGFR4 enzyme solution was added to a 384-well plate, and the final concentration of the enzyme was 0.1 5 nM. 2) 1 5 μL of diluted solution in gradient of the compound was added. 3) 1 5 μL of a substrate mixture containing substrate polypeptide with a final concentration of 5 50 nM and ATP with a final concentration of 10 200 μM was added. 4) The mixture was incubated for 0.5 3 hours at room temperature. 5) 10 μL of EDTA and a test solution comprising labeled antibody were added, and the plate was incubated for 1 hour at room temperature. 6) The fluorescence signal values of each plate were determined by a microplate reader at 665 nm. 7) The inhibition rates were calculated according to the fluorescence signal values. |
| Affinity data for this assay | |
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