Unknown

HEK 293T cells transfect with CB1 or CB2 receptor were used for activity evaluation. Forty-eight hours after transfection, the cells were washed twice with Opti-MEM I reduced serum medium to remove any remaining FBS, and 100 μL of Opti-MEM I was added. The Nano-Glo Live Cell reagent, a nonlytic detection reagent containing the cellpermeable furimazine substrate, was prepared by diluting the Nano-Glo Live Cell substrate 20× using Nano-Glo LCS Dilution buffer, and 25 μL was added to each well. Subsequently, the plate was placed in a GloMAX96 (Promega). Luminescence was monitored during the equilibration period until the signal was stabilized (30−45 min). For agonist experiments, we added 10 μL per well of test compounds, present as 13.5× stocks in 50% methanol in Opti-MEM I. Luminescence was continuously detected for 120 min. The luminescence was continuously detected for 120 min.