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HTS046M ChemiSCREEN™ Membrane Preparation Recombinant Human ETB Endothelin Receptor

Human ETB GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.

MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
Host Cell: CHO (Chinese hamster ovary cells) Uni Prot Number: P24530 Entrez Gene Number: NM_00015
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HTS046M
200 units  
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      Overview

      Key Spec Table

      SpeciesHost CellsProtein Target
      Human CHO (Chinese hamster ovary cells)ETB
      Description
      Catalogue NumberHTS046M
      Brand Family Chemicon®
      Trade Name
      • ChemiScreen
      • Chemicon
      DescriptionChemiSCREEN™ Membrane Preparation Recombinant Human ETB Endothelin Receptor
      OverviewHuman ETB isoform 1
      Background InformationThe endothelin family of peptides are potent vasoconstrictors synthesized by endothelial cells in both constitutive and inducible pathways. The biological actions of endothelins are mediated by two GPCRs, ETA and ETB (Davenport, 2002). Whereas ETA is the predominant receptor on smooth muscle cells and thus is the primary mediator of the vasoconstrictor activity, ETB is found on endothelial cells and mediates vasodilation (Nilsson et al., 1997). In addition, ETB has also been linked to renal failure, congestive heart failure, atherosclerosis, and pulmonary hypertension (D’Orléans-Juste, et al., 2002). A mutation in ETB has been found to cause Hirschsprung disease-2, a degenerative disease of the digestive tract (Puffenberger et al, 1994). Millipore’s cloned human ETB expressing cell line is made in the CHO host, which supports high levels of recombinant ETB expression on the cell surface. Thus, the ETB Membrane Preparation is an ideal tool for screening for compounds binding to ETB. The membrane preparations exhibit a Kd of 0.03-0.06 nM for [125I]-endothelin-1. With 0.2 nM [125I]-endothelin-1, 10 μg/well ETB Membrane Prep yields greater than 5 fold signal-to-background ratio.
      Product Information
      FormatMembranes
      PresentationOne package contains enough membranes for at least 200 assays (units), where an unit is the amount of membrane that will yield greater than 5-fold signal:background with 125I-labeled Endothelin-1 at 0.25 nM.

      Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA with no preservatives.
      Packaging method: Membrane protein was adjusted to the indicated concentration in packaging buffer, rapidly frozen, and stored at -80°C.
      Quality LevelMQ100
      Applications
      ApplicationHuman ETB GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.
      Key Applications
      • Radioligand binding assay
      • GTPγS binding
      Application NotesRadioligand binding assay and GTPγS binding
      Biological Information
      Entrez Gene Number
      Gene Symbol
      • EDNRA
      • ETA
      • ETRA
      GPCR ClassA
      Host CellsCHO (Chinese hamster ovary cells)
      Protein TargetETB
      Target Sub-FamilyEndothelin
      UniProt Number
      Incubation ConditionsRECOMMENDED 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 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]Endothelin-1 (Perkin Elmer # NEX259)

      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 an unit is the amount of membrane that will yield greater than 5-fold signal:background with 125I-labeled Endothelin-1 at 0.25 nM.
      Table 1. Signal:background and specific binding values obtained in a competition binding assay with ETB membrane prep.

      Product Usage Statements
      10 µg/well
      Signal:Background 7.4
      Specific Binding (cpm) 5704


      SPECIFICATIONS: 1 unit = 10 μg membrane preparation
      Bmax: 0.49 pmol/mg
      Kd: 0.03-0.06 nM 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.
      Storage and Shipping Information
      Storage ConditionsStore at –70°C. Product is stable for at least 6 months from the date of receipt when stored as directed. Do not freeze and thaw.
      Packaging Information
      Material Size200 units
      Global Trade Item Number
      Catalogue Number GTIN
      HTS046M 04053252273209