| Assay Method Information | |
| | CB1 receptor binding assay |
| Description: | Mouse brain membranes were used as the source material for CB1 receptors. The displacement of specifically bound tritiated CP-55.940 from these membranes using a standard filtration assay was used to determine the Ki values for the test compounds. Briefly. 20 μg of protein was incubated for 1 h at 30° C. in the presence of 0.5 nM [3H]CP-55,940 and various concentrations (10−5M-10−11M) of test compound/control, final volume of 1 mL. The incubation was terminated by rapid filtration and washing, and the amount of specifically bound [3H]CP-55,940 was determined. Briefly, membranes with bound [3H]CP-55,940 were separated and washed from free ligand by vacuum filtration, adsorbing the membrane onto a Whatman glass microfiber filter paper (LIFEGENE, Cat #1821271). Finally, the Whatman filter paper with adsorbed membranes was cut and placed in scintillation liquid (Ultima Gold) for 1 h at 25° C. followed by a β counter reading of bound [3H]CP-55,940 radioligand. All data were in triplicates with Ki values determined using GraphPad Prism 7.02 analysis software. Data normalized between 0 and 100% specific binding were plotted against log concentration of test compound, and Ki was extracted using nonlinear regression analysis. |
| Affinity data for this assay | |
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