| Assay Method Information | |
| | Assessing the Affinity of the Compounds of the Present Disclosure for the 5-HT2A Receptor |
| Description: | Method: The affinity of the compounds of the present disclosure for the 5-HT2A receptor was determined using a radioligand competitive binding assay.In the first step, a cell membrane component containing 5-HT2A receptor was prepared as follows. HEK-293T cells (ATCC, CRL-11268) in a 10-cm dish were transfected with 10 ng of 5-HT2A receptor plasmid and 40 μL of PEI. The 10-cm dish was removed from the incubator 48 hours post-transfection, a time point by which the cultured cells had expressed 5. HT2A receptor protein. The culture medium was aspirated using a vacuum pump, and 3 mL of lysis buffer was added to each dish. The cells were then placed in a cold room at 4° C. for 10 minutes. After the cells were detached, they were transferred to a 15 mL centrifuge tube and centrifuged at 1500 rpm for 5 minutes at 4° C., and the supernatant was discarded. The cell pellet was transferred to a tissue homogenizer, and 3 mL of lysis buffer was added. Sufficient grinding was applied to break the cells. The cell suspension was then equally aliquoted into several Eppendorf tubes and centrifuged at 12,000 rpm for 5 minutes at 4° C., and the supernatant was discarded. The resulting precipitate was the cell membrane component containing the 5-HT2A receptor. |
| Affinity data for this assay | |
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