| Assay Method Information | |
| | Assay on Agonistic Effect of Compounds on β1 Receptor by HTRF cAMP Method |
| Description: | β1 cells (expressing human ADRB1 gene) from ICE Bioscience Inc. were grown under standard conditions. The cells were collected and diluted with 1 Stimulation Buffer, and 9 μL of the cell dilution was added to a white low-volume 384-well plate with 4000 cells seeded in each well. Compounds were serially diluted in DMSO in a 5-fold gradient to obtain 10 concentrations. Before testing, compounds that had been serially diluted in DMSO were subjected to 100-fold dilution with 1 Stimulation Buffer to obtain compound working solutions. The concentration of the vehicle DMSO was 0.1%. Eu-cAMP and ULight -anti-cAMP were diluted to the working concentration with Detection buffer. 5 μL of Eu-cAMP diluent and 5 μL of ULight -anti-cAMP diluent were sequentially added into corresponding experiment wells. After 1 h of incubation at room temperature, readings at 665 nm and 620 nm were measured under the excitation wavelength of 330 nm using a Biotek microplate reader. The activity of the compounds was obtained by plotting through Ratio (665/620) against the concentration of the compounds, fitting the curve by a nonlinear regression method using GraphPad Prism 7 |
| Affinity data for this assay | |
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