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Figure 1. Saturation binding for Y4. 5 µg/well Y4 Membrane Preparation was incubated with increasing amount of 125I-labeled Pa...
Figure 2. Competition binding for Y4 . 5µg/well Y4 Membrane Preparation and wild-type Chem-1 Membrane Preparation (Chemicon catalog # HTS000MC1) were...
ChemiSCREEN™ Membrane Preparation Recombinant Human Y4 Neuropeptide Y Receptor
Description:
ChemiSCREEN™ Membrane Preparation Recombinant Human Y4 Neuropeptide Y Receptor
Trade Name:
- ChemiScreen
- Chemicon (Millipore)
Qty/Pk:
200 units
Product Overview:
Full-length human PPYR1 cDNA encoding Y4
Background Information:
The NPY family consists of three 36-amino acid peptides, neuropeptide Y (NPY), peptide YY (PYY) and pancreatic polypeptide (PP), which bind to the NPY receptor family of G protein-coupled receptors. Five NPY receptors, Y1, Y2, Y4, Y5 and Y6, have been defined at the molecular level, and each signals primarily through Gi/o. Binding of NPY family peptides to NPY receptors mediates a variety of physiological effects, including promotion of food intake, decreased anxiety, inhibition of neurotransmitter and hormone release, vasoconstriction, and gut motility. Y4 binds preferentially to PP, with significant binding to NPY and PYY (Michel et al., 1998). Y4 plays a role in control of reproduction, as Y4-null, ob/ob double knockout mice have increased fertility relative to ob/ob mice with functional Y4 (Sainsbury et al., 2002). In addition, mice lacking Y4 display resistance to weight gain induced by high fat diet, and mice lacking both Y2 and Y4 are resistant to the enhanced weight gain exhibited by Y1-null mice fed a high fat diet (Sainsbury et al., 2006). Millipore's Y4 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 Y4. The membrane preparations exhibit a Kd of 0.35 nM for [125I]-Pancreatic Polypeptide. With 0.4 nM [125I]-Pancreatic Polypeptide, 5μg/well Y4 Membrane Prep typically yields greater than 20-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.
Species:
Human
Quality Assurance:
Table 1. Signal:background and specific binding values obtained in a competition binding assay with 5 μg/well of Y4 Receptor membrane prep.
SPECIFICATIONS: 1 unit = 5 µg
Bmax for [125I]-Pancreatic Polypeptide binding: 3.7 pmol/mg protein
Kd for [125I]-Pancreatic Polypeptide binding: ~0.35 nM
| 5 µg/well | |
|---|---|
| Signal:Background | 37.5 |
| Specific Binding (cpm) | 9558 |
SPECIFICATIONS: 1 unit = 5 µg
Bmax for [125I]-Pancreatic Polypeptide binding: 3.7 pmol/mg protein
Kd for [125I]-Pancreatic Polypeptide binding: ~0.35 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, 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: 50 mM Hepes, pH 7.4, 5 mM MgCl2, 1mM CaCl2, 0.2% BSA filtered and stored at 4°C
Radioligand: [125I]-Pancreatic Polypeptide. (Perkin Elmer #: NEX315 )
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 20-fold signal:background with 125I labeled Pancreatic Polypeptide at 0.4 nM.
Binding buffer: 50 mM Hepes, pH 7.4, 5 mM MgCl2, 1mM CaCl2, 0.2% BSA filtered and stored at 4°C
Radioligand: [125I]-Pancreatic Polypeptide. (Perkin Elmer #: NEX315 )
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 20-fold signal:background with 125I labeled Pancreatic Polypeptide at 0.4 nM.
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 the indicated concentration in 1 ml packaging buffer, rapidly frozen, and stored at -80°C.
Packaging method: Membranes protein were adjusted to the indicated concentration in 1 ml packaging buffer, rapidly frozen, and stored at -80°C.
Protein Target:
Y4
Ligand Type:
peptide
Storage Conditions:
Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.
Target Sub-Family:
Neuropeptide Y
Promotional Text:
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GPCR Class:
A
Protein or Enzyme Type:
GPCR Membrane Preparations
Product Name:
ChemiSCREEN™ Membrane Preparation Recombinant Human Y4 Neuropeptide Y Receptor
Entrez Gene Number:
Cell Membrane Type:
GPCR