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HTS104M

Sigma-Aldrich

ChemiSCREEN Membrane Preparation Recombinant Human β1 Adrenoceptor Receptor

Human beta 1 GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.

Synonym(s):

β₁ Adrenoceptor Membrane Prep, ChemiSCREEN Receptor Kit

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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

manufacturer/tradename

ChemiScreen
Chemicon®

technique(s)

ligand binding assay: suitable (GTPγS)
radioligand binding assay (RLBA): suitable

NCBI accession no.

UniProt accession no.

shipped in

dry ice

Gene Information

human ... ADRB1(153)

General description

Full-length human ADRB1 cDNA encoding the β1 adrenoceptor
The endogenous catecholamines epinephrine and norepinephrine have profound effects on smooth muscle activity, cardiac function, carbohydrate and fat metabolism, hormone secretion, neurotransmitter release, and central nervous system actions. These activities are mediated by GPCRs belonging to two subfamilies, the α- and β-adrenergic receptors (Bylund et al., 1994). The three members of the β-adrenergic receptor family, β1, β2 and β3, couple to Gs to increase cAMP upon activation. In the heart, the β1 receptor constitutes 70-80% of the β-adrenergic receptors. Activation of cardiac β-adrenergic receptors acutely increases heart rate, cardiac output, and cardiac automaticity, and chronically increases cardiac myocyte apoptosis. In failing hearts, the β1 subtype is downregulated and desensitized, probably as a result of increased catecholamine levels. As a result, β-adrenergic receptor antagonists (β blockers) are effective in the treatment of congestive heart failure and arrhythmia (Lohse et al., 2003). Millipore′s β1 adrenoceptor 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 β1 adrenoceptor interactions. The membrane preparations exhibit a Kd of 2.6 nM for [125I]-(-) Iodocyanopindolol (ICYP). With 5 µg/well β1 Adrenoceptor Membrane Prep and 0.25 nM [125I]-(-)ICYP, a greater than 15-fold signal-to-background ratio was obtained.

Application

Radioligand binding assay and GTPγS binding.

Biochem/physiol Actions

GPCR Class: A
Protein Target: beta 1
Target Sub-Family: Adrenergic

Quality

5 µg/well 10 µg/well
Signal:Background 28.5 34.6
Specific Binding (cpm) 27002 28467


SPECIFICATIONS: 1 unit = 5 μg membrane preparation
Bmax: 47.9 pmol/mg
Kd: ~2.6 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, 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. 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, filtered and stored at 4°C
Radioligand: [125I]-(-) Iodocyanopindolol (Perkin Elmer # NEX189)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl, filtered and stored at 4°C.

Physical form

One package contains enough membranes for at least 200 assays (units), where an unit is the amount of membrane that will yield greater than 15-fold signal:background with 125I-labeled (-)Iodocyanopindolol at 0.25 nM.

Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA with no preservatives.
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 Code

12 - Non Combustible Liquids

WGK

WGK 2

Flash Point(F)

Not applicable

Flash Point(C)

Not applicable


Certificates of Analysis (COA)

Search for Certificates of Analysis (COA) by entering the products Lot/Batch Number. Lot and Batch Numbers can be found on a product’s label following the words ‘Lot’ or ‘Batch’.

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What is the role of beta-adrenergic signaling in heart failure?
Lohse, Martin J, et al.
Circulation Research, 93, 896-906 (2003)

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