CHEMBL5734680

Receptor Binding Assay: Histamine H1: (1) The prepared membrane was applied with appropriate amount of buffer, and homogenizer was used for evenly dispersing. 15 tubes were mixed into a 100 ml container, and appropriate amount of buffer (potassium dihydrogen phosphate 1.36 g, 0.1 mol/L sodium hydroxide 79 ml, metered to 200 ml with double-distilled water) was added to give 50 ml of membrane suspension, which was reserved for future use. (2) 100 uL of membrane preparation was added into each reaction tube. (3) 100 uL of buffer was added into the total binding tube (TB), 100 uL of promethazine (final concentration 10-5M) was added into the nonspecific binding tube (NB), 100 uL of the test compound (final concentration 10-5M) was added into the specific binding tube (SB) of each test compound. (4) 10 uL of radioactive ligand 3H-pyrilamine was respectively added into each reaction tube (2 parallel tubes were used for each reaction tube, and each of them was placed on ice when adding sample). (5) Each of the reaction tubes was incubated at 30° C. for 60 min. After the reaction was completed, the bound ligands were rapidly filtered under reduced pressure, and the ice-chilled assay buffer was used for adequate washing. The filter was taken out and put into a 3 ml scintillation vial, and 2 ml of toluene scintillation solution was added and blended. 2* Isotope ligand [3H]-5-CT (85.4 Ci/mmol) was purchased from PerkinElmer Company; (+-)-pindolol was purchased from RBI Company; GF/C glass fiber filter paper was purchased from Whatman Company; Tris was imported and divided into aliquots; PPO and POPOP were purchased from Shanghai No. 1 Reagent Factory; liposoluble scintillation cocktail was purchased from Shanghai Reagent Factory. Beckman LS-6500 Multi-function Liquid Scintillation Counter was used.