ChemiScreen BLT1 Membrane Preparation

32161000

NA.84

BLT1 receptor is a Gi-coupled GPCR that binds primarily to the eicosanoid leukotriene B4. Macrophages, leukocytes and eosinophils express BLT1, and in vivo, BLT1 appears to mediate leukocyte migration and inflammation (reviewed in Brink et al., 2003). Chemicon′s BLT1 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 BLT1 interactions with Leukotriene B4. The membrane preparations exhibit a Kd of 1.3 nM for [3H]-Leukotriene B4. With 5.0 μg/well BLT1 Membrane Prep and 1 nM [3H]-Leukotriene B4, a signal-to-background ratio of approximately 3-fold is obtained.
Human BLT1

Radioligand binding assay, and GTPgammaS binding.
Human BLT1 GPCR membrane preparation for Radioligand binding Assays & GTPgammaS binding.

Target Sub-Family: Leukotriene
GPCR Class: A
Protein Target: BLT1

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

	5 µg/well	10 µg/well
Signal:Background	75	65
Specific Binding (cpm)	46348	54457

1 unit = 5µg membrane preparation
Bmax: 5.6 pmol/mg
Kd: 0.43 nM

Inucbation Conditions
RECOMMENDED 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, 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] Luekotriene (Perkin Elmer# NEX378)

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 2.5-fold signal:background or 500 cpm with 3H-labeled Leukotriene B4 at 1 nM

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

Packaging method: Membranes protein were adjusted to the indicated concentration in packaging buffer, rapidly frozen, and stored at -80°C.

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

100

human

expressed in Chem-1 cells

pkg of 1 mL

ChemiScreen
Chemicon^®

0.5 mg/mL (per vial)

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

NM_181657.1

Q15722

dry ice

Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.

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.

12 - Non Combustible Liquids

WGK 2