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Figure 1. Saturation binding for EP2. 10 µg/well EP2 Membrane Preparation was incubated with increasing amount of 3
Figure 2. Competition binding for EP2. 10 µg/well EP2 Membrane Preparation and wild-type Chem-1 Membrane Preparation (Chemicon ...
ChemiSCREEN™ Membrane Preparation Recombinant Human EP2 Prostanoid Receptor
Description:
ChemiSCREEN™ Membrane Preparation Recombinant Human EP2 Prostanoid Receptor
Trade Name:
Chemicon (Millipore)
Qty/Pk:
200 units
Product Overview:
Full-length human PTGER2 cDNA encoding EP2
Background Information:
Prostanoids bind to a family of 8 GPCRs to exert their biological effects (Narumiya and FitzGerald, 2001). The protanoid PGE2 causes pain, vasodilation, immunosuppression of T cells, bone resorption and promotion of carcinogenesis. Four related GPCRs, EP1, EP2, EP3 and EP4, each bind to PGE2, but the different G protein coupling status of each receptor leads to distinct biological effects. EP2 couples primarily to Gs to increase intracellular cAMP levels. Mice deficient in EP2 receptor showed impaired ovulation and fertilization, salt-sensitive hypertension (Kennedy et al., 1999). It has been shown that EP2 receptors are also involved in cancer associated immunodeficiency. Thus, genetic knockout of the EP2 receptor reduced tumor growth and prolonged survival in mice that had undergone isograft injection of MC26 or Lewis lung carcinoma cells (Yang et al., 2003). Millipore's EP2 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 agonists and antagonists of EP2. The membrane preparations exhibit a Kd of 14.52 nM for [3H]-PGE2. With 7.5 nM [3H]-PGE2, 10µg/well EP2 Membrane Prep typically yields greater than 3-fold signal-to-background ratio.
Application Notes:
Radioligand binding assay and GTPγS binding
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.
UniProt Summary:
FUNCTION: SwissProt: P43116 # Receptor for prostaglandin E2 (PGE2). The activity of this receptor is mediated by G(s) proteins that stimulate adenylate cyclase. The subsequent raise in intracellular cAMP is responsible for the relaxing effect of this receptor on smooth muscle.
SIZE: 358 amino acids; 39761 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
TISSUE SPECIFICITY: Placenta and lung.
SIMILARITY: SwissProt: P43116 ## Belongs to the G-protein coupled receptor 1 family.
SIZE: 358 amino acids; 39761 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
TISSUE SPECIFICITY: Placenta and lung.
SIMILARITY: SwissProt: P43116 ## Belongs to the G-protein coupled receptor 1 family.
Species:
Human
Quality Assurance:
Table 1. Signal:background and specific binding values obtained in a competition binding assay with varying amounts of EP2 Receptor membrane prep.
1 unit = 10 µg
Bmax for [3H]-PGE2 binding: 4.38 pmol/mg protein
Kd for [3H]-PGE2 binding: ~14.52 nM
| 10 µg/well | |
|---|---|
| Signal:Background | 3.60 |
| Specific Binding (cpm) | 1146 |
1 unit = 10 µg
Bmax for [3H]-PGE2 binding: 4.38 pmol/mg protein
Kd for [3H]-PGE2 binding: ~14.52 nM
Cell Type:
Chem-1
Brand Family:
Chemicon
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, 500mM NaCl. 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, filtered and stored at 4°C.
Radioligand: [3H]-PGE2 (Perkin Elmer#: NET-428)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl, filtered and stored at 4°C.
Binding buffer: 50 mM Hepes, pH 7.4, 5 mM MgCl2, 1 mM CaCl2, filtered and stored at 4°C.
Radioligand: [3H]-PGE2 (Perkin Elmer#: NET-428)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl, filtered and stored at 4°C.
Host Cells:
Chem-1
Presentation:
Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA with no preservatives.
Packaging method: Membranes protein were adjusted to 1 mg/mL in packaging buffer, and dispensed at 1 mL/vial. Vials were rapidly frozen, and stored at -80°C.
Packaging method: Membranes protein were adjusted to 1 mg/mL in packaging buffer, and dispensed at 1 mL/vial. Vials were rapidly frozen, and stored at -80°C.
Protein Target:
EP2
UniProt Number:
Ligand Type:
non-peptide
Storage Conditions:
Store at –70°C. Product is stable for at least 6 months from the date of receipt when stored as directed. Do not freeze and thaw.
Target Sub-Family:
Prostanoid
Promotional Text:
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GPCR Class:
A
Gene Symbol:
- PTGER2
- EP2
Protein or Enzyme Type:
GPCR Membrane Preparations
Product Name:
ChemiSCREEN™ Membrane Preparation Recombinant Human EP2 Prostanoid Receptor
Entrez Gene Number:
Cell Membrane Type:
GPCR