ChemiSCREEN Membrane Preparation Recombinant Human BB₃Bombesin Receptor

半岛bd体育手机客户端 说明

一般描述

Full length human BRS3 encoding BB₃
Bombesin, a bioactive peptide first identified in amphibian skin, is related to two mammalian peptides, gastrin-releasing peptide (GRP) and neuromedin B. A family of 3 GPCRs, including GRP-R (BB₁), NMB-R (BB₂) and BRS-3 (BB₃), mediate the biological effects of the peptides (Ohki-Hamazaki et al., 2005). BB₃differs from the others by its low affinity for bombesin. Although an endogenous ligand for BB₃has yet to be identified, a synthetic nonselective bombesin-like peptide [H-D-Phe6, β-Ala11, Phe13, Nle14]-bombesin-(6-14)-nonapeptide amide (Bombesin (6-14) Analog) activates BB₃with high potency. BB₃-null mice have an obese phenotype (Matsumoto and Iijima, 2003). Millipore′s BB₃membrane 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 BB₃receptor interactions with its ligand. The membrane preparations exhibit a Kd of 0.23nM for [125I]-Bombesin (6-14) Analog. With 10 μg/well BB₃Membrane Prep and 0.3 nM [¹²⁵I]-Bombesin (6-14) Analog, a greater than 3-fold signal-to-background ratio was obtained.

应用

Human BB3 GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.
Radioligand binding assay and GTPγS binding

生化/生理作用

• GPCR Class: A
• Protein Target: BB3
• Target Sub-Family: Bombesin

半岛bd体育手机客户端 规格

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, 1 mM CaCl₂, 0.2% BSA, 0.2 mg/ml bacitracin, 20 μg/ml leupeptin, 20 μg/ml chymostatin, 1 Protease Inhibitor cocktail Tablets (Roche Cat. No. 11 873 580 001) for each 50 ml binding buffer.

Radioligand: [¹²⁵I]-Bombesin (6-14) Analog (Perkin Elmer NEX377)

Wash Buffer: 50 mM Hepes, pH 7.4, 500 mM NaCl , 0.1% BSA, filtered and stored at 4℃.

One package contains enough membranes for at least 200 assays (units), where an unit is the amount of membrane that will yield greater than 3-fold signal:background with¹²⁵I-labeled Bombesin (6-14) Analog at 0.3 nM.

外形

Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA with no preservatives.
Packaging method: Membranes protein was adjusted to the indicated concentration in packaging buffer, rapidly frozen, and stored at -80oC.

储存及稳定性

Maintain frozen at -70℃ for up to 2 years. Do not freeze and thaw.

法律信息

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

免责声明

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

基本信息

eCl@ss

32161000

半岛bd体育手机客户端 性质

质量水平	100
生物来源	human
重组	expressed in Chem-1 cells
manufacturer/tradename	ChemiScreen Chemicon?
technique(s)	ligand binding assay: suitable (GTPγS) radioligand binding assay (RLBA): suitable
NCBI登记号	NM_001727
UniProt登记号	P32247

安全信息

储存分类代码	12 - Non Combustible Liquids
WGK	WGK 2
闪点(F)	Not applicable
闪点(C)	Not applicable