Cp55 940

Manufactured by Bio-Techne

Sourced in United Kingdom, United States

CP55,940 is a high-affinity cannabinoid receptor agonist. It binds to both CB1 and CB2 receptors with high potency.

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Lab products found in correlation

Am251, by Bio-Techne (6 mentions) Win55 212 2, by Bio-Techne (4 mentions) Penicillin, by Merck Group (3 mentions) Streptomycin, by Merck Group (3 mentions) 35s gtpγs, by PerkinElmer (3 mentions) Am630, by Bio-Techne (3 mentions) Bovine serum albumin (bsa), by Merck Group (3 mentions) 35s gtpγs, by American Radiolabeled Chemicals (2 mentions) Damgo, by Bio-Techne (2 mentions) Trizmatm hydrochloride, by Merck Group (2 mentions) Microscinttm 20, by PerkinElmer (2 mentions) Non enzymatic cell dissociation solution, by Merck Group (2 mentions) Dimethyl sulfoxide (dmso), by Merck Group (2 mentions) Urb602, by Cayman Chemical (2 mentions) O 1602, by Cayman Chemical (1 mentions) Anti rhoa 26c4 mouse monoclonal antibody, by Santa Cruz Biotechnology (1 mentions) Lipofectaminetm 2000, by Thermo Fisher Scientific (1 mentions) Y 27632, by Bio-Techne (1 mentions) Sr141716a, by Bio-Techne (1 mentions) Jwh 018, by Bio-Techne (1 mentions)

28 protocols using cp55 940

Regulation of Lipid Signaling Pathways

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Arachidonic acid (20:4,n-6), essentially fatty acid-free bovine serum albumin (BSA), and lysophosphatidic acid (LPA) (1-oleoyl) sodium salt were obtained from Sigma (St. Louis, MO, USA). 1,2-Dipalmitoyl PI and 1,2-dioleoyl PI were from Avanti Polar Lipids, Inc. (Alabaster, AL, USA).
CP55940 and Y-27632 were from Tocris (Bristol, UK). WIN55212-2 was obtained from RBI (Natick, MA, USA). O-1602 was obtained from Cayman Chemical Co. (Ann Arbor, MI, USA). PD98059, SB203580, and SP600215 were acquired from Calbiochem-Novabiochem (San Diego, CA, USA). Wortmannin was obtained from Wako Pure Chem. Ind. (Osaka, Japan). Clostridium botulinum C3 exoenzyme pcDNA4/TO and Lipofectamine^TM 2000 were from Invitrogen Life Technologies (Carlsbad, CA, USA). The anti-RhoA 26C4 mouse monoclonal antibody was from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). The anti-mouse IgG horseradish-peroxidase-linked goat antibody was obtained from Medical & Biological Laboratories Co, Ltd. (Nagoya, Japan). Lipase (Rhizopus delemar) was acquired from Seikagaku Kogyo Co., Ltd. (Tokyo, Japan).

Nakajima K., Oka S., Tanikawa T., Nemoto-Sasaki Y., Matsumoto N., Ishiguro H., Arata Y., Sugiura T, & Yamashita A. (2022). Lysophosphatidylinositol Induced Morphological Changes and Stress Fiber Formation through the GPR55-RhoA-ROCK Pathway. International Journal of Molecular Sciences, 23(18), 10932.

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Cannabinoid Compound Preparation

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2-AG, AEA, and CP55,940 were purchased from Tocris Bioscience (Bristol, U.K.). Org27569, PSNCBAM-1, and GAT100 were synthesized in the laboratory of Dr. Ganesh A Thakur (Northeastern University, Boston, MA). All compounds were dissolved in DMSO (final concentration, 0.1% in culture media for all cell-based assays) and added directly to the media at concentrations and times indicated.

Laprairie R.B., Kulkarni A.R., Kulkarni P.M., Hurst D.P., Lynch D., Reggio P.H., Janero D.R., Pertwee R.G., Stevenson L.A., Kelly M.E., Denovan-Wrigh E.M, & Thakur G.A. (2016). Mapping Cannabinoid 1 Receptor Allosteric Site(s): Critical Molecular Determinant and Signaling Profile of GAT100, a Novel, Potent, and Irreversibly Binding Probe. ACS chemical neuroscience, 7(6), 776-798.

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Cannabinoid Compounds Characterization

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CP55,940 and SR141716A were purchased from Tocris Bioscience (Oakville, ON). All other cannabinoids were obtained at ≥ 98% purity from Aurora Prairie (Aurora Cannabis Inc., Saskatoon, SK). Because concern exists regarding the stability plant-derived cannabinoids, such as ∆⁹-THCa undergoing spontaneous decarboxylation, all compounds were aliquoted, stored at − 80 °C until use, and were used only once. Compounds were assessed for purity by high performance liquid chromatography with diode-array detection (HPLC-DAD) using well-described methods following both purification and 1-month storage at − 80 °C^{57 (link)}. A representative chromatogram for ∆⁹-THCa is included in Supplementary Figure S1. [³H]CP55,940 (174.6 Ci/mmol) was obtained from PerkinElmer (Guelph, ON). All other reagents were obtained from Sigma-Aldrich (Oakville, ON) unless specifically noted. Compounds were dissolved in DMSO (final concentration of 0.1% in assay media for all assays) and added directly to the media at the concentrations and times indicated.

Zagzoog A., Mohamed K.A., Kim H.J., Kim E.D., Frank C.S., Black T., Jadhav P.D., Holbrook L.A, & Laprairie R.B. (2020). In vitro and in vivo pharmacological activity of minor cannabinoids isolated from Cannabis sativa. Scientific Reports, 10, 20405.

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Synthesis and Assay of Cannabinoid Receptor Ligands

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IQD analogues PNR-4-20 and PNR-4-02 were synthesized as previously described (30 (link)) and were dissolved in 100% DMSO to 10 mM stock concentrations for in vitro assays. WIN-55,212-2, JWH-018, CP-55,940, and DAMGO were obtained from Tocris Bioscience (Bristol, United Kingdom). GTPγS was purchased from EMD Chemical (Gibbstown, NJ). [³H]CP-55,940 (52.6 Ci/mmol) was acquired from PerkinElmer (Waltham, MA) and [³⁵S]GTPγS (1250 Ci/mmol) was obtained from American Radiolabeled Chemicals (St. Louis, MO). PathHunter™ enzyme fragment complementation (EFC) experimental reagent was purchased from DiscoverRx Corporation (Fremont, CA). All other reagents used were purchased from Fisher Scientific (Pittsburgh, PA).

Ford B.M., Franks L.N., Tai S., Fantegrossi W.E., Stahl E.L., Berquist M.D., Cabanlong C.V., Wilson C.D., Penthala N.R., Crooks P.A, & Prather P.L. (2017). Characterization of Structurally Novel G Protein Biased CB1 Agonists: Implications for Drug Development. Pharmacological research, 125(Pt B), 161-177.

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Purification and Characterization of Membrane Proteins

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Oligonucleotides were purchased from Operon Biosciences. Restriction enzymes and DNA-modifying enzymes were obtained from New England Biolabs. The Complete His resin was from Roche. The purified Strep-Tactin XT, Strep-Tactin XT Superflow and MagStrep “type3” XT beads were kindly provided by IBA GmbH. Research grade sensor chip CM4, immobilization reagents NHS, EDC, and ethanolamine and HBS-N buffers for SPR experiments were from GE Healthcare.
Cholesteryl hemisuccinate Tris salt (CHS) and detergents 3 [(cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS) and n-dodecyl-b-D-maltosyde (DDM) were obtained from Anatrace. Lipids 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine sodium salt (POPS) were purchased from Avanti Polar Lipids Inc. Synthetic cannabinoid ligand CP-55,940 was from Tocris. All other chemicals of reagent grade were purchased from Sigma.

Yeliseev A., Zoubak L, & Schmidt T.G. (2016). Application of Strep-Tactin XT for affinity purification of Twin-Strep-tagged CB2, a G protein-coupled cannabinoid receptor. Protein expression and purification, 131, 109-118.

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Cannabinoid Receptor Binding Assay

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CP55,940 was purchased from Tocris Bioscience (Minneapolis, MN, USA). BSA, Trizma^TM hydrochloride (Tris-HCl), penicillin and streptomycin, nonenzymatic cell dissociation solution, and guanosine 5′-diphosphate (GDP) were purchased from Sigma-Aldrich (St. Louis, MO, USA). [³H]-CP55,940 and MicroScint^TM-20 were purchased from PerkinElmer (Waltham, MA, USA). Membrane preparation was made using a 50 mM Tris-HCl buffer with pH 7.4. Dilutions of membrane, radioligand and control/test compounds were made in a Tris-EDTA buffer (50 mM Tris-HCl, 20 mM EDTA, 154 mM NaCl, and 0.2% fatty-acid BSA), with pH = 7.4.

Pandey P., Roy K.K., Liu H., Ma G., Pettaway S., Alsharif W.F., Gadepalli R.S., Rimoldi J.M., McCurdy C.R., Cutler S.J, & Doerksen R.J. (2018). Structure-Based Identification of Potent Natural Product Chemotypes as Cannabinoid Receptor 1 Inverse Agonists. Molecules : A Journal of Synthetic Chemistry and Natural Product Chemistry, 23(10), 2630.

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Pharmacological Modulation of Neuronal Cytoskeleton

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CB1R agonists WIN55,212-2, CP-55940, HU-210, 2-arachidonoylglycerol (2-AG), and CB1R-specific antagonist/inverse agonists AM281 and AM251 were acquired from Tocris Bioscience (Bristol, UK). Rho-associated kinase inhibitor Y-27632 and nocodazole were purchased from Calbiochem (San Diego, CA). Blebbistatin, cytochalasin D, ML-7, Δ⁹-Tetrahydrocannabidiol solution (THC), and pertussis toxin (PTX) were brought from Sigma (Saint-Louis, MO). The Rho-GTPase inhibitor C3 transferase (C3T) was purchased from Cytoskeleton, Inc. Mouse anti-neuron-specific beta III tubulin (Tuj-1) antibody was obtained from Sigma (Catalog Number T8660), rabbit anti-myosin phospho S19/phospho S20 antibody was obtained from Rockland (Gilbertsville, PA, Cat. no. 600-401-416), rabbit anti-CB1R antibody was produced by Eurogentec (Seraing, Belgium) and described previously (Thibault et al., 2013 (link)), and chicken anti-GFP antibody was from AVES (Tigard, OR). Alexa-Fluor-conjugated secondary antibodies were purchased from Life Technologies (Carlsbad, CA). All culture media and additives were from PAA Laboratories (Pasching, Austria).

Roland A.B., Ricobaraza A., Carrel D., Jordan B.M., Rico F., Simon A., Humbert-Claude M., Ferrier J., McFadden M.H., Scheuring S, & Lenkei Z. (2014). Cannabinoid-induced actomyosin contractility shapes neuronal morphology and growth. eLife, 3, e03159.

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Ligand Binding and Cell Signaling Assay

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G418 was purchased from KSE Scientific (Durham, NC). Puromycin, DMEM, penicillin/streptomycin, gentamicin, DPBS, Hank’s Buffer, HEPES, EDTA, Tris base, sucrose, MgCl₂, Millipore filter plates and punch kits, EcoLite scintillation cocktail, and Poly-d-lysine coated 96-well plates were purchased from Fisher Scientific (Waltham, MA). Ambisome and FBS were purchased from Atlanta Biologicals (Flowery Branch, GA). Ro 20-1724, acetonitrile, DMSO, lipopolysaccharide (LPS), polyethyleneamine, and fatty acid-free BSA were purchased from Sigma Aldrich (St. Louis, MO). Antibodies against murine CD16/32 (ab33550) and CD206 (ab64693) were purchased from Abcam (Cambridge, UK). High bind plates (L15XB-3), anti-rat (R32AA-5), and anti-rabbit (R32AB-1) SULFO-TAG antibodies were purchased from Meso-Scale Discovery (Gaithersburg, MD). Cellular dyes phalloidin and Hoechst were purchased from Cellomics (Pittsburgh, PA), and ACTOne Membrane Potential Dye was purchased from Codex BioSolutions (Gaithersburg, MD). Forskolin, CP 55,940, HU 308, JWH 133, and SR 144528 were purchased from Tocris (Bristol, UK). Tritiated CP 55,940 was purchased from PerkinElmer (Waltham, MA). The 96-well filter plates and punch tips were purchased from Millipore (Billerica, MA).

Presley C., Abidi A., Suryawanshi S., Mustafa S., Meibohm B, & Moore B.M. (2015). Preclinical evaluation of SMM-189, a cannabinoid receptor 2-specific inverse agonist. Pharmacology Research & Perspectives, 3(4), e00159.

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Cannabinoid Receptor Binding Assay

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CP55940 [(-)-cis-3-[2-Hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl)cyclohexanol] was purchased from Tocris Bioscience (Bristol, United Kingdom). AEA and indomethacin were purchased from Sigma-Aldrich (Poole, Dorset, United Kingdom). [³H]CP55940 (174.6 Ci/mmol) and [³⁵S]GTPγS (1250 Ci/mmol) were obtained from PerkinElmer (Seer Green, Buckinghamshire, United Kingdom), GTPγS from Roche Diagnostic (Burgess Hill, West Sussex, United Kingdom), and GDP from Sigma-Aldrich. Compounds were dissolved in DMSO (final concentration of 0.1% in assay media for all assays) and added directly to the media at the concentrations and times indicated.

Laprairie R.B., Mohamed K.A., Zagzoog A., Kelly M.E., Stevenson L.A., Pertwee R., Denovan-Wright E.M, & Thakur G.A. (2019). Indomethacin Enhances Type 1 Cannabinoid Receptor Signaling. Frontiers in Molecular Neuroscience, 12, 257.

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Radioligand Binding Assay for CB1 Receptor

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Approximately 3 μg of membrane preparation expressing CB₁ was incubated with nine concentrations of allosteric modulator (1 nM-10 μM). The radiolabeled tracer [^{3 (link)}H]CP55,940 (150.2 Ci/mmol; Perkin Elmer), an orthosteric agonist of CB₁, was added at 0.5 nM. Nonspecific binding was established by treating with a high concentration of unlabeled CP55,940 (10 μM; Tocris). The membranes were incubated at 30°C for 60 min, and the reaction was terminated with the addition of 300 μL of Tris-Mg²⁺-EDTA (TME) buffer with 5% bovine serum albumin (BSA). The mixture was harvested by filtration through a Brandel cell harvester with Whatman GF/C filter paper. Liquid scintillation counting was used to measure radioactivity.

Immadi S.S., Dopart R., Wu Z., Fu B., Kendall D.A, & Lu D. (2018). Exploring 6-Azaindole and 7-Azaindole Rings for Developing Cannabinoid Receptor 1 Allosteric Modulators. Cannabis and Cannabinoid Research, 3(1), 252-258.

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