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由来生物
human
品質水準
リコンビナント
expressed in Chem-1 cells
メーカー／製品名
ChemiScreen
Chemicon®
テクニック
ligand binding assay: suitable (GTPγS)
radioligand binding assay (RLBA): suitable
NCBIアクセッション番号
UniProtアクセッション番号
輸送温度
dry ice
詳細
Full-length human CHRM3 cDNA encoding M3
The muscarinic acetylcholine receptor (mAChR) family consists of five GPCRs that mediate some of the neurotransmission functions of acetylcholine in the CNS and the periphery. The M1, M3 and M5 receptors couple to Gq to mobilize intracellular calcium, whereas the M2 and M4 receptors couple to Gi/o to inhibit cAMP production (Caulfield and Birdsall, 1998). M3 is expressed prominently in smooth muscle, and plays a primary role in mediating mAChR agonist-induced contractility. Mice lacking M3 have dilated pupils, which indicates a role for M3 in regulating tone of the pupillary sphincter muscle. In addition, Msub3 plays a role in feeding, as indicated by the lean and hypophagic phenotype of M3-null mice (Wess, 2004). Chemicon′s M3 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 M3 interactions with 4-DAMP. The membrane preparations exhibit a Kd of 0.72-1 nM for [3H]-4-DAMP. With 10 µg/well M3 Membrane Prep and 0.75 nM [3H]-4-DAMP, a greater than 4-fold signal-to-background ratio was obtained.
アプリケーション
Human M3 GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.
Radioligand binding assay and GTPγS binding.
生物化学的／生理学的作用
GPCR Class: A
Protein Target: M3
Target Sub-Family: Acetylcholine (muscarinic)
品質
Signal:background and specific binding values obtained in a competition binding assay with varying amounts of M3 membrane prep:
| 20 µg/well | 10 µg/well | |
|---|---|---|
| Signal:Background | 5.17 | 5.10 |
| Specific Binding (cpm) | 1224 | 1030 |
SPECIFICATIONS: 1 unit = 10 µg
Bmax: 3.75 pmol/mg
Kd: 0.86 nM
規格
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, 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: [3H]-4-DAMP (Perkin Elmer#:NET1040 )
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 a unit is the amount of membrane that will yield greater than 4-fold signal:background with 3H labeled 4-DAMP at 0.75 nM
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: [3H]-4-DAMP (Perkin Elmer#:NET1040 )
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 a unit is the amount of membrane that will yield greater than 4-fold signal:background with 3H labeled 4-DAMP at 0.75 nM
物理的形状
Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA no preservatives.
Packaging method: Membranes protein were adjusted to 0.5 mg/ml in 1 mL packaging buffer, rapidly frozen, and stored at -80°C.
Packaging method: Membranes protein were adjusted to 0.5 mg/ml in 1 mL packaging buffer, rapidly frozen, and stored at -80°C.
保管および安定性
Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.
法的情報
CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany
免責事項
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.
保管分類コード
12 - Non Combustible Liquids
WGK
WGK 2
引火点(°F)
Not applicable
引火点(℃)
Not applicable
適用法令
試験研究用途を考慮した関連法令を主に挙げております。化学物質以外については、一部の情報のみ提供しています。 製品を安全かつ合法的に使用することは、使用者の義務です。最新情報により修正される場合があります。WEBの反映には時間を要することがあるため、適宜SDSをご参照ください。
Jan Code
HTS116M:
試験成績書（COA）
製品のロット番号・バッチ番号を入力して、試験成績書（COA） を検索できます。ロット番号・バッチ番号は、製品ラベルに「Lot」または「Batch」に続いて記載されています。
International Union of Pharmacology. XVII. Classification of muscarinic acetylcholine receptors.
M P Caulfield et al.
Pharmacological reviews, 50(2), 279-290 (1998-07-02)
Muscarinic acetylcholine receptor knockout mice: novel phenotypes and clinical implications.
Wess, Jurgen
Annual Review of Pharmacology and Toxicology, 44, 423-450 (2004)
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