| Assay Method Information | |
| | Radioligand Binding Assay |
| Description: | For the membrane binding, NIH-3T3 cells were grown to 70% confluence in 15-cm2 dishes and transfected with 10 ug of receptor plasmid DNA using Polyfect transfection reagent (QIAGEN, Valencia, CA) according to manufacturer s protocols. Two days after transfection, cells expressing the desired serotonin receptor were homogenized in 20 mM HEPES/10 mM EDTA and spun down at 11,000g at 4 C for 30 min. The supernatant was discarded, and the pellet was resuspended in 20 mM HEPES/1 mM EDTA and spun down at the same setting. The pellet was then resuspended in 20 mM HEPES/0.5 mM EDTA, and membranes were used for binding assays. Bradford analysis was used to determine total membrane protein. |
| Affinity data for this assay | |
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