Key Specifications Table
| Species | Host Cells | Protein Target |
|---|---|---|
| Human | Chem-1 | DP |
| Description | |
|---|---|
| Catalogue Number | HTS091M |
| Brand Family | Chemicon® |
| Trade Name |
|
| Description | ChemiSCREEN™ Human DP Prostanoid Receptor Membrane Preparation |
| Overview | Full-length human PTGDR cDNA encoding DP |
| Background Information | Prostanoids are a series of arachidonic acid metabolites produced by the action of cyclooxygenase and subsequently by isomerases and synthases. Cells rapidly secrete prostanoids after synthesis, whereupon the prostanoids bind to a family of 8 GPCRs to exert their biological effects (Narumiya and FitzGerald, 2001). The prostaglandin PGD2 is produced by mast cells upon activation by allergens, and is present at high levels in allergic diseases. PGD2 binds to two receptors, DP and CRTH2. DP activates Gs to increase cAMP levels, and lack of DP results in reduced allergic response in animal models of bronchial asthma (Matsuoka et al., 2000). Chemicons DP 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 DP interactions and its ligands. The membrane preparations exhibit a Kd of 10.39 nM for [3H]-PGD2. With 8 nM [3H]-PGD2, 5 µg/well and 10 µg/well DP Membrane Prep typically yield greater than 4-fold signal-to-background ratio. |
| Product Information | |
|---|---|
| Format | Membranes |
| Presentation | 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. |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | Human DP GPCR membrane preparation for Radioligand binding Assays & GTPgammaS binding. |
| Key Applications |
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| Application Notes | Radioligand binding assay and GTPS binding. |
| Biological Information | |
|---|---|
| Entrez Gene Number |
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| Entrez Gene Summary | The protein encoded by this gene is a G-protein-coupled receptor. It has been shown to function as a prostanoid DP receptor. The activity of this receptor is mainly mediated by G-S proteins that stimulate adenylate cyclase resulting in an elevation of intracellular cAMP and Ca2+. Knockout studies in mice suggest that the ligand of this receptor, prostaglandin D2 (PGD2), functions as a mast cell-derived mediator to trigger asthmatic responses. |
| Gene Symbol |
|
| GPCR Class | A |
| Host Cells | Chem-1 |
| Protein Target | DP |
| Target Sub-Family | Prostanoid |
| UniProt Number |
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| UniProt Summary | FUNCTION: SwissProt: Q13258 # Receptor for prostaglandin D2 (PGD2). The activity of this receptor is mainly mediated by G(s) proteins that stimulate adenylate cyclase, resulting in an elevation of intracellular cAMP. A mobilization of calcium is also observed, but without formation of inositol 1,4,5-trisphosphate. SIZE: 359 amino acids; 40271 Da SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein. TISSUE SPECIFICITY: Expressed in retina and small intestine. DISEASE:SwissProt: Q13258 # Genetic variations in PTGDR are associated with susceptibility to asthma-related traits type 1 (ASRT1) [MIM:607277]. Asthma-related traits include clinical symptoms of asthma, such as coughing, wheezing and dyspnea. SIMILARITY: SwissProt: Q13258 ## Belongs to the G-protein coupled receptor 1 family. |
| Incubation Conditions | 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, a GF/C 96-well filter plate 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: 50 mM Hepes, pH 7.4, 5 mM MgCl2, 1 mM CaCl2, 0.2% BSA, filtered and stored at 4°C Radioligand: [3H] PGD2 (Perkin Elmer#:NET-616 ) Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl , 0.1% BSA, filtered and stored at 4°C. One package contains enough membranes for at least 200 assays (units), where a unit is the amount of membrane that will yield greater than 4-fold signal:background with 3H-labeled PGD2 at 8 nM |
| Product Usage Statements |
|---|
| 10 µg/well | 5 µg/well | |
|---|---|---|
| Signal:Background | 4.86 | 4.12 |
| Specific Binding (cpm) | 1345 | 885 |
Table 2. IC50 values for ligands obtained in a competition binding assay with DP membrane preparation.
| IC50 (nM) | |
|---|---|
| Prostaglanding D2 | 12.1 |
| 13,14-dihydro-15-keto prostaglandin D2 | >10000 |
| BW A868C | 4.8 |
SPECIFICATIONS: 1 unit = 10 µg
Bmax for [3H] PGD2 binding: 5.3 pmol/mg protein;
Kd for [3H] PGD2 binding: ~10.39nM Usage Statement
- 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.
| Storage and Shipping Information | |
|---|---|
| Storage Conditions | Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw. |
| Packaging Information | |
|---|---|
| Material Size | 200 units |