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| | TR-FRET Farnesoid X Receptor Coactivator Assay |
| Description: | Purchasing invitrogen PV4833 kits.First, the required amount of the compound was weighed and dissolved in 100% DMSO at the maximum concentration of 3000 μM. The solution at the maximum concentration was diluted by 3-fold serial dilution in DMSO to get 10 concentrations;Second, the above prepared solutions of different concentrations were diluted to 100-fold with a buffer supplied with the kit, followed by mixing and then 10 μL of the diluted solution was added to a 384 well plate;Third, nuclear receptor FXR recombinant protein was diluted with a buffer to give a concentration of 4×, and 5 μL of the diluent was added to the 384 well plate of second step;Fourth, Fluorescein-SRC2-2 and Tb anti-GST antibody were diluted with a buffer to give a concentration of 4× respectively. Then two reagents were mixed together, and 10 μL of the mixture was added to the 384 well plate of third step;Finally, the solution of the 384 well above was mixed uniformly by centrifuging, and then incubated at room temperature for 1 h. Then the TR-FRET Endpoint was used for measuring the solution at wavelengths of 520 nm, 495 nm and 337 nm. EC50 values were calculated according to the measured value of ER=520 nm/495 nm. |
| Affinity data for this assay | |
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