ChemiSCREEN Membrane Preparation Recombinant Human CCK₁Cholecystokinin Receptor

半岛bd体育手机客户端 说明

一般描述

Human CCK₁Receptor
Cholecystokinins are a series of peptides of heterogeneous length (5 to 58 amino acids) that are derived from preprocholecystokinin and are found in gastrointestinal tissues and the central nervous system. Gastrin is a related peptide with 5 C-terminal amino acids identical to those of cholecystokinin. Two GPCRs, CCK₁(CCK_A) and CCK₂(CCK_B), bind to CCK and/or gastrin to mediate the biological effects of the peptides. CCK₁selectively binds sulfated CCK, whereas CCK₂binds to CCK and gastrin with similar affinity. Binding of ligands to CCK₁stimulates mobilization of intracellular calcium by activation of G_q/11. CCK₁receptors in the periphery are primarily localized in the pancreas, gallbladder, pylorus, intestine where they are responsible for the regulation of diverse digestive processes. They are also present in select areas of the peripheral nervous system (vagus nerve), and the CNS where they mediate the satiety effects of CCK, regulate an increase in dopamine release, and antagonize opioid analgesia (Noble et al., 1999). Chemicon′s CCK₁membrane 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 CCK₁interactions and its ligands. The membrane preparations exhibit a Kd of 0.5 nM for [¹²⁵I]-CCK-8 sulfated. With 10.0ug/well CCK1 membrane prep and 0.5 nM [¹²⁵I]-CCK-8 sulfated, a greater than 4-fold signal-to-background ratio was obtained.

应用

Radioligand binding assay and GTPγS binding

生化/生理作用

Protein Target: CCK1 / CCKa
GPCR Class: A
Target Sub-Family: Cholecystokinin

质量

Table 1. Signal:background and specific binding values obtained in a competition binding assay with varying amounts of CCK₁Receptor membrane prep.

	10 ?g/well	5 ?g/well
Signal:Background	5.31	6.16
Specific Binding (cpm)	1274.9	1691.4

SPECIFICATIONS: 1 unit = 10 ?g membrane preparation
Bmax: 0.15 pmol/mg
K _d: 0.5 nM

半岛bd体育手机客户端 规格

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, an FC 96-well harvest plate (Millipore cat. # MAHF C1H) is blocked 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.
Note: Due to the acidic property of the labeled sulfated CCK-8, filter plates coated with PEI bind the labeled ligand nonspecifically and result in elevated backgrounds.

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] -CCK-8 sulfated (Perkin Elmer # NEX203)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl, 0.1% BSA, filtered and stored at 4°C.

外形

One package contains enough membranes for at least 200 assays (units), where an unit is the amount of membrane that will yield greater than 4-fold signal:background with¹²⁵I-labeled CCK-8 at 0.5 nM.

Liquid in packaging buffer: 50 mM Tris, pH 7.4, 10% glycerol 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°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
NACRES	NA.41

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_000730.2
UniProt登记号	P32238
运输	dry ice

安全信息

储存分类代码	12 - Non Combustible Liquids
WGK	WGK 1
闪点(F)	Not applicable
闪点(C)	Not applicable