Chemoattractant receptor homologous molecule expressed on Th2 cells (CRTH2) is a GPCR related to FPR-like chemoattractant receptors that is highly expressed on Th2 cells, eosinophils, basophils, and type 2 cytotoxic T cells (Nagata et al., 1999). The most potent ligand for CRTH2 is the prostaglandin PGD2, a major mast cell-derived allergic mediator (Hirai et al., 2001). Another GPCR, DP, is activated by PGD2, and the relative contributions of DP and CRTH2 to PGD2-mediated allergic disease are being elucidated. A small molecule antagonist of CRTH2, ramatroban, attenuates PGD2-mediated bronchial hyperresponsiveness and antigen-induced inflammation (Sugimoto et al., 2003; Shichijo et al., 2003).Chemicon′s CRTH2 membrane preparations are crude membrane preparations made from our proprietary stable recombinant cell lines to ensure high-level of GPCR surface expression; thus, they are ideal HTS tools for screening of antagonists of PGD2/CRTH2 interactions. The membrane preparations exhibit a Kd of 10 nM for [3H]-Prostaglandin D2. With 2.5 μg/well CRTH2 Membrane Prep and 10 nM [3H]-Prostaglandin D2, a greater than 5-fold signal-to-background ratio is obtained.
200 units in 1 mL
Radioligand binding assay, and GTPgammaS binding.
GPCR Class: A
Target Sub-Family: Prostanoid
Protein Target: CRTH2 / DP2 / GPR44
Signal:background and specific binding values obtained in a competition binding assay with CRTH2 membrane preparation with unlabeled Prostaglandin D2.
Specific Binding (cpm) 4307
IC50 values for ligands obtained in a competition binding assay with CRTH2 membrane preparation.
IC50 (nM) Prostaglanding D2 4.5
13,14-dihydro-15-keto prostaglandin D2 14.7
BW A868C 7429
SPECIFICATIONS: 1 unit = 10 g
Bmax: 13.93 pmol/mg
Kd: 10 nM
Membranes are mixed with radioactive ligand and unlabeled competitor (see Figures 1 and 2 for concentrations tested) in binding buffer in a nonbinding 96-well plate, and incubated for 1-2 h. Prior to filtration, an FC 96-well harvest plate (Millipore cat. # MAHF C1H) is coated with 0.33% polyethyleneimine for 30 min, then washed with 50mM HEPES, pH 7.4, 0.5% BSA. Binding reaction is transferred to the filter plate, and washed 3 times (1 mL per well per wash) with Wash Buffer. The plate is dried and counted.
Binding buffer: 50mM HEPES, 10mM MnCl2, 1mM EDTA, 0.2% BSA, pH to 7.0 and filter.
Wash buffer: 50mM HEPES, 0.5M NaCl, pH to 7.4 and filter.
Radioligand: [3H] Prostaglandin D2 (Amersham#: TRK734)
Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA no preservatives.Packaging method: Membranes protein were adjusted to 0.5 mg/ml in 1 ml packaging buffer, rapidly frozen, and stored at -80°C.
Stockage et stabilité
Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.
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Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.