ChemiSCREEN™ Membrane Preparation Recombinant Human CB₂ Canabinoid Receptor Optimized for GTPγS Assay

ChemiSCREEN™ Membrane Preparation Recombinant Human CB₂ Canabinoid Receptor Optimized for GTPγS Assay

• ChemiScreen
• Chemicon (Millipore)

200 units

Human CB₂

Cannabinoid compounds include exogenous drugs such as Δ⁹-THC, the main psychoactive component of the plant Cannabis sativa, and endogenous mediators, such as anandamide, that belong to eicosanoid family.The biological effects of cannabinoids are mediated by a family of two Gi-coupled 7-transmembrane receptors, CB₁ and CB₂. The CB₁ receptor is found primarily in brain and mediates the psychoactive effects of cannabinoid ligands. The CB₂ receptor is expressed mainly in immune cells, including mast cells and CD40-activated B cells, where it mediates proliferation and inhibition of migration (Howlett et al., 2002). Activation of CB₂ inhibits the development of liver fibrosis (Julien et al., 2005). In bone, CB₂ is expressed in both osteoblasts and osteoclasts, and functions to prevent bone loss (Ofek et al., 2006). In addition, activation of CB₂ has an antinociceptive effect in animal models of neuropathic, inflammatory, and acute pain; this effect is mediated by release of endogenous opioids in the periphery (Ibrahim et al., 2005).
Millipore's CB₂ Membrane Preparation for GTPγS Assay is a crude membrane preparation made from the proprietary stable recombinant cell line that is also used for Millipore’s CB₂ Calcium Optimized Cell Line (HTS020C). This CB₂ Membrane Preparation has been optimized for low basal activity in GTPγS binding assays; thus it is an ideal HTS tool for screening of compounds for agonist and antagonist activities. The membrane preparations exhibits EC50 of 13.7 nM for CP55,940 in a GTPγS binding assay. This CB₂ Membrane Preparation may also be used in radioligand binding assays.
Millipore offers another CB₂ Membrane Preparation (HTS020M) which has been optimized for high signal:background in radioligand binding assay, and also displays high constitutive activity that permits analysis of compounds for inverse agonist activity.

Radioligand binding assay and GTPγS binding

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.

Human

SPECIFICATIONS: 1 unit = 5 μg
EC50 in GTPγS binding assay by CP55940: ~ 13.7 nM
Bmax: 23.6 pmol/mg; Kd: 3.1 nM

Chem-4

Chemicon

Membranes are permeabilized by addition of saponin to an equal concentration by mass, then mixed with [³⁵S]-GTPγS (final concentration of 0.3 nM) in 20 mM HEPES, pH 7.4/100 mM NaCl/10 mM MgCl₂/10 μM GDP in a nonbinding 96-well plate. Unlabeled CP55940 was added to the final concentration indicated in Figure 1 (final volume 100 μL), and incubated for 30 min at 30°C. The binding reaction is transferred to an FB filter plate (Millipore MAHF B1H) previously prewetted with water. The plate is washed 3 times (1 mL per well per wash) with cold 10 mM sodium phosphate, pH 7.4, then dried and counted.

One vial contains enough membranes for at least 200 assays (units), where one unit is the amount of membrane that will yield greater than 1000 cpm specific CP55940-stimulated [35S]-GTPγS binding.

Chem-4

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.

CB2

non-peptide

Store 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

Cannabinoid

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A

GPCR Membrane Preparations

ChemiSCREEN™ Membrane Preparation Recombinant Human CB₂ Canabinoid Receptor Optimized for GTPγS Assay

X74328

GPCR