Cyclosporin A, Inhibitor

Cyclosporin A

Cat. Num. AAA575587

• Purity: 99.9%
• Synonyms: Cyclosporin A; inhibitor

• Purity/Purification: 99.9%
• Form/Format: POWDER

• CAS: 59865-13-3
• Formula: C62H111N11O12
• Solubility: DMSO: 120.3 mg/mL (100 mM); Ethanol: 60.1 mg/mL (50 mM)
• Receptor (IC50): calcineurin phosphatase: 7nM
• Kinase Assay: Phosphatase Assay: Purified bovine brain calcineurin and calmodulin are purchased. Reaction mixtures with purified enzyme contains 100 nM calcineurin, 100 nM calmodulin, and 5 uM 32P-labeled phosphopeptide, in 60 ul (total volume) of assay buffer containing 20 mM Tris (pH 8), 100 mM NaCI, 6 mM MgCl2, 0.5 mM dithiothreitol, 0.1 mg of bovine serum albumin per ml, and either 0.1 mM CaCl2 or 5 mM EGTA. Reaction mixtures with cell lysates contains 20 ul of undiluted lysate, 5 uM 32P-labeled phosphopeptide, and 40 ul of assay buffer. Where indicated, reaction mixtures contains 50 uM peptide 412 or 413 and/or 500 nM okadaic acid, a specific inhibitor of phosphatases 1 and 2A; 500 nM okadaic acid is sufficient for inhibition of Ca2+-independent phosphatases, whereas higher concentrations partially inhibit Ca2+-dependent activity as well. After 15 min at 30°C,reactions are terminated by the addition of 0.5 ml of 100 mM potassium phosphate buffer (pH 7.0) containing 5% trichloroacetic acid. Free inorganic phosphate is isolated by Dowex cation-exchange chromatography and quantitated by scintillation counting as described.
• Cell Assay: Immunosuppressive agents are dissolved in ethanol at concentrations 1000-fold more than the concentration desired for cell treatments. Cells (106) are suspended in 1 ml of complete medium in microcentrifuge tubes; 1 ul of ethanol or of the ethanolic solution of Cyclosporin A is added, and the cells are incubated at 37°C for 1 hr. Cells are washed twice with 1 ml of PBS on ice and lysed in 50ul of hypotonic buffer containing 50 mM Tris (pH 7.5); 0.1 mM EGTA; 1 mM EDTA; 0.5 mM dithiothreitol; and 50 ug of phenylmethylsulfonyl fluoride, 50 ug of soybean trypsin inhibitor, 5 ug of leupeptin, and 5 ug of aprotinin per ml. Lysates are subjected to three cycles of freezing in liquid nitrogen followed by thawing at 30°C and then are centrifuged at 4°C for 10 min at 12,000×g. (Only for Reference)
• Animal Experiment: Animal Model: Rat
• Smiles: CC[C@@H]1NC(=O)[C@H]([C@H](O)[C@H](C)C\C=C\C)N(C)C(=O)[C@H](C(C)C)N(C)C(=O)[C@H](CC(C)C)N(C)C(=O)[C@H](CC(C)C)N(C)C(=O)[C@@H](C)NC(=O)[C@H](C)NC(=O)[C@H](CC(C)C)N(C)C(=O)[C@@H](NC(=O)[C@H](CC(C)C)N(C)C(=O)CN(C)C1=O)C(C)C
• Preparation and Storage: 2 years -80°C in solvent; 3 years -20°C powder

Structure