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Figure 1. Saturation binding for VPAC1. 5 µg/well VPAC1 Membrane Preparation was incubated with increasing amount of 125I-labe ...
Figure 2. Competition binding for VPAC1. 5 and 10 µg/well VPAC1 Membrane Preparation (Chemicon cat. # HTS043M) and 10 µg/well ...
ChemiScreen™ VPAC1 Membrane Preparation
Description:
ChemiScreen™ VPAC1 Membrane Preparation
Trade Name:
- ChemiScreen
- Chemicon (Millipore)
Product Overview:
Human VIPR1 cDNA, encoding VPAC1
Background Information:
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 Notes:
Radioligand binding assay, and GTPgammaS 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.
Cell Membrane Type:
GPCR
Descriptive Text:
Concentration:
1 mL, 0.5 mg/mL per vial.
Host Cells:
Chem-1, an adherent mammalian cell line without any endogenous VPAC1 expression.
Cell Type:
Chem-1
Gene Symbol:
- VIPR1
- HVR1
- PACAP-R-2
- RCD1
- VPCAP1R
- II
- FLJ41949
- VIP-R-1
- VAPC1
- VPAC1
- VIPR
- VIRG
- RDC1
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 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.
Quality Assurance:
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
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: [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
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
Protein or Enzyme Type:
GPCR Membrane Preparations
Storage Conditions:
Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.
Species:
Human