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Phoenix Amphotropic (AMPHO) cell line 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 (Promega), a nonlytic detection reagent containing the cell permeable 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 luminometer, the GloMAX96 (Promega). Luminescence was monitored during the equilibration period until the signal stabilized (30−45 min). For agonist experiments, we added 10 μL per well of test compounds (UR-144 N-pentanoic acid), present as 13.5× stocks in 50% methanol in Opti-MEM I. The luminescence was continuously detected for 120 min. Solvent controls were run in all experiments; the final concentration of methanol (3.7%) did not pose a problem given the short readout time of the assay.