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HTS090M Sigma-Aldrich

ChemiScreen Recombinant Human OT Oxytocin Receptor Membrane Preparation

Human OT GPCR membrane prepation for Radioligand binding Assays.

  •  eCl@ss 32161000

  •  NACRES NA.84

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Properties

Quality Level   100
biological source   human
recombinant   expressed in Chem-1 cells
mfr. no.   ChemiScreen
  Chemicon®
application(s)   radioligand binding assay (RLBA): suitable
NCBI accession no.   NM_000916.3
UniProt accession no.   P30559
shipped in   dry ice
Gene Information   human ... OXTR(5021)

Description

General description

Human Oxytocin Receptor

Oxytocin is a cyclic 9 amino acid peptide that differs from vasopressin in 2 amino acids. Despite the close similarities in sequence, oxytocin and vasopressin have different biological activities and bind to distinct G protein-coupled receptors. The oxytocin receptor, OT, couples primarily to Gq/11 to mobilize intracellular calcium. In female reproduction, oxytocin promotes uterine contraction and lactation; oxytocin is the most commonly used drug for induction of labor, whereas an oxytocin antagonist, atosiban, is under investigation to suppress preterm labor. Oxytocin/OT interaction in the CNS also plays an important role in stress, male and female sexual response, and sociality (Gimpl and Fahrenholz, 2001). Chemicon′s oxytocin 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 oxytocin receptor interactions with oxytocin. The membrane preparations exhibit a Kd of 0.53nM for [125I]-OVTA. With 5.0 ug/well Oxytocin Membrane Prep and 0.5 nM [125I]-OVTA, a greater than 6-fold signal-to-background ratio was obtained.

Biochem/physiol Actions

GPCR Class: A

Protein Target: OT

Target Sub-Family: Oxytocin

Quality

Signal:background and specific binding values obtained in a competition binding assay with varying amounts of Oxytocin Receptor membrane prep:

5 µg/well10 µg/well
Signal:Background 9 9
Specific Binding (cpm) 21871 24926


SPECIFICATIONS: 1 unit = 5 µg membrane preparation
Bmax: 4.5 pmol/mg
Kd: 0.53 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, a GF/C 96-well filter plate 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] OVTA (Perkin Elmer # NEX254)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl , 0.1% BSA, 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 6-fold signal:background with 125I-labeled OVTA at 0.5 nM.

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 -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.

Safety & Documentation

Safety Information

WGK Germany 
WGK 2
Flash Point(F) 
Not applicable
Flash Point(C) 
Not applicable
Protocols & Articles
Peer-Reviewed Papers
15

References

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