ChemiSCREEN Membrane Preparation Recombinant Human 5-HT_1BSerotonin Receptor

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一般描述

Full-length human HTR1B cDNA encoding the 5-HT_1BSerotonin Receptor
5-Hydroxytryptamine (5-HT, also commonly known as serotonin) is synthesized in enterochromaffin cells in the intestine and in serotonergic nerve terminals. In the periphery, 5-HT mediates gastrointestinal motility, platelet aggregation, and contraction of blood vessels. Many functions of the central nervous system are influenced by 5-HT, including sleep, motor activity, sensory perception, arousal and appetite. A family of 12 GPCRs and one ion channel mediate the biological effects of 5-HT (Hoyer et al., 1994). The 5-HT_1Breceptor (also known as 5-HT_1Dβ) is expressed presynaptically on serotonergic neurons and postsynaptically on non-serotonergic neurons, and it regulates release of 5-HT and acetylcholine. Several brain regions, including globus pallidus, substantia nigra and dorsal subiculum, highly express HT1B. 5-HT_1Bhas been implicated in the physiology of aggression, depression, migraine, anxiety and reward for drugs of abuse (Sari, 2004). 5-HT_1Binteracts with p11, which increases cell surface expression and function of 5-HT_1B(Svenningsson et al., 2006). Millipore′s 5-HT_1Bmembrane 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 5-HT_1B. The membrane preparations exhibit a Kd of 10.5 nM for [³H]-GR125743. With 7 nM [³H]-GR125743, 10μg/well 5-HT_1BMembrane Prep typically yields greater than 8-fold signal-to-background ratio.

应用

Radioligand binding assay and GTPγS binding.

生化/生理作用

Target Sub-Family: Serotonin
GPCR Class: A
Protein Target: 5-HT1B

质量

Table 1. Signal:background and specific binding values obtained in a competition binding assay with varying amounts of sst5 Receptor membrane prep.

	10 ?g/well	20 ?g/well
Signal:Background	12.4	10.5
Specific Binding (cpm)	3210	3352

SPECIFICATIONS: 1 unit =10 μg
Bmax for [ ³H]-GR125743 binding: 3.7 pmol/mg protein
Kd for [ ³H]-GR125743 binding: ~10.5 nM

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Inucbation Conditions
RECOMMENDED ASSAY 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 Tris, pH 7.4. 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 Tris, pH 7.4, 10 mM MgCl2, 1 mM EDTA, filtered and stored at 4°C

Radioligand: [³H]-GR125743. (Amersham #:TRK-1046 )

Wash Buffer: 50 mM Tris, pH 7.4, 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 8-fold signal:background with³H labeled GR125743 at 7 nM

外形

Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA no preservatives.
Packaging method: Membranes protein were adjusted to 1 mg/ml in 1 ml packaging buffer, rapidly frozen, and stored at -80°C.

储存及稳定性

Maintain frozen at -70°C 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 uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

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_000863.1
UniProt登记号	P28222
运输	dry ice

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