HTS043M
ChemiScreen VPAC1 Membrane Preparation
Human VPAC1 / VIP1 GPCR membrane preparation for Radioligand binding Assays & GTPgammaS binding.
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About This Item
UNSPSC Code:
41106514
eCl@ss:
32161000
NACRES:
NA.84
biological source
human
Quality Level
recombinant
expressed in Chem-1 cells
packaging
pkg of 1 mL
manufacturer/tradename
ChemiScreen
Chemicon®
concentration
0.5 mg/mL (per vial)
technique(s)
ligand binding assay: suitable (GTPγS)
radioligand binding assay (RLBA): suitable
NCBI accession no.
UniProt accession no.
shipped in
dry ice
General description
Human VIPR1 cDNA, encoding VPAC1
Vasoactive intestinal peptide (VIP), a 28 amino acid peptide originally isolated by its vasodilation activity, binds to two class B GPCRs, VPAC1 and VPAC2, to exert its functions in the CNS, vasculature, immune system and adrenal medulla (Harmar et al., 1998). In the immune system, VIP is synthesized by mast cells and lymphocytes, and appears to inhibit inflammation and to shift the immune response toward a Th2 pathway (Delgado et al., 2004). In the heart, VIP is expressed by nerve fibers, where it modulates heart rate, and coronary blood flow (Henning and Sawmiller, 2001). Chemicon′s VPAC1 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 VPAC1 interactions with VIP. The membrane preparations exhibit a Kd of 1.2 nM for [125I]-VIP. With 2.5 μg/well VPAC1 Membrane Prep and 0.4 nM [125I]-VIP, a greater than 5-fold signal-to-background ratio is obtained.
Application
Radioligand binding assay, and GTPgammaS binding.
Biochem/physiol Actions
GPCR Class: B
Protein Target: VPAC1 / VIP1
Target Sub-Family: VIP/PACAP
Quality
Signal:background and specific binding values obtained in a competition binding assay with varying amounts of VPAC1 membrane prep:
SPECIFICATIONS:
1 unit = 5 µg
Bmax: 19.65 pmol/mg
Kd: 1.2 nM
| 10 µg/well | 5 µg/well | |
|---|---|---|
| Signal:Background | 9.7 | 10.8 |
| Specific Binding (cpm) | 30299 | 31224 |
SPECIFICATIONS:
1 unit = 5 µg
Bmax: 19.65 pmol/mg
Kd: 1.2 nM
Specifications
Inucbation 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: [125I] VIP (Perkin Elmer# NEX192)
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 5-fold signal:background with 125I-labeled VIP at 0.4 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, 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: [125I] VIP (Perkin Elmer# NEX192)
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 5-fold signal:background with 125I-labeled VIP at 0.4 nM
Physical form
Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA no preservatives.
Packaging method: Membranes protein were adjusted to the indicated concentration in packaging buffer, rapidly frozen, and stored at -80°C.
Packaging method: Membranes protein were adjusted to the indicated concentration in packaging buffer, rapidly frozen, and stored at -80°C.
Storage and Stability
Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.
Legal Information
CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany
Disclaimer
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 Class
12 - Non Combustible Liquids
wgk_germany
WGK 2
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Find documentation for the products that you have recently purchased in the Document Library.
International Union of Pharmacology. XVIII. Nomenclature of receptors for vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide.
A J Harmar et al.
Pharmacological reviews, 50(2), 265-270 (1998-07-02)
Mario Delgado et al.
Pharmacological reviews, 56(2), 249-290 (2004-06-01)
First identified by Said and Mutt some 30 years ago, the vasoactive intestinal peptide (VIP) was originally isolated as a vasodilator peptide. Subsequently, its biochemistry was elucidated, and within the 1st decade, their signature features as a neuropeptide became consolidated.
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