| Assay Method Information | |
| | Binding Assay |
| Description: | A whole cell receptor binding assay using [3H]ramatroban as the competing radioactive ligand was employed to evaluate the compound binding activity to human CRTH2. The radioactive ligand [3H]ramatroban was synthesized according to Sugimoto et. al. (Eur. J. Pharmacol. 524, 30-37, 2005) to a specific activity of 42 Ci/mmol.A cell line stably expressing human CRTH2 was established by transfecting CHO-K1 cells with two mammalian expression vectors that harbored human CRTH2 and G-alpha16 cDNAs, respectively, using FuGene 6 transfection reagent (from Roche). Stable clones expressing CRTH2 were selected by staining each clone with BM16 (BD Pharmingen from BD Biosciences, a division of Becton, Dickinson and Company), which is a rat monoclonal antibody to human CRTH2. The cells were maintained as monolayer cultures in Ham's F-12 medium containing 10% fetal bovine serum, 100 units/mL penicillin, 100 ug/mL streptomycin, 2 mM glutamine, 0.5 mg/mL G418 (geneticin) for CRTH2. |
| Affinity data for this assay | |
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