Structure
CHIR-99021, inhibitor
CHIR-99021
Purity
98.95%
Synonyms
CHIR-99021; inhibitor
Purity/Purification
98.95%
Form/Format
POWDER
CAS
252917-06-9
Formula
C22H18Cl2N8
Biological Activity
GSK-3Alpha/Beta inhibitor (IC50: 10/6.7 nM).
Solubility
DMSO: 9.3 mg/mL
Receptor (IC50)
GSK-3Alpha: 10 nM (cell free)
GSK-3Beta: 6.7 nM (cell free)
GSK-3Beta: 6.7 nM (cell free)
Smiles
Cc1cnc([nH]1)-c1cnc(NCCNc2ccc(cn2)C#N)nc1-c1ccc(Cl)cc1Cl
Preparation and Storage
Store at -20 degree C.
Structure
Structure
Related Product Information for CHIR-99021, inhibitor
Kinase Assay
Kinases were purified from SF9 cells through the use of their His or Glu tag. Glutagged proteins were purified as described, and His-tagged proteins were purified according to the manufacturer's instructions. Kinase assays were performed in 96-well plates with appropriate peptide substrates in a 300-1J1 reaction buffer (variations on 50 mM Tris-HCI, pH 7.5, 10 mM MgCl2, 1 mM EGTA, 1 mM dithiothreitol, 25 mM~-glycerophosphate, 1 mM NaF, and 0.01% bovine serum albumin). Peptides had Km values from 1 to 100 uM. CHIR 99021 or CHIR GSKIA was added in 3.5 ul of Me2SO, followed by ATP to a final concentration of 11JM. After incubation, triplicate 100-u1 aliquots were transferred to Combiplate 8 plates containing 100 ul/well of 50 uM ATP and 20 mM EDTA. After 1 hour, the wells were rinsed five times with phosphate-buffered saline, filled with 200 IJI of scintillation fluid, sealed, and counted in a scintillation counter 30 min later. All of the steps were at room temperature. The percentage of inhibition was calculated as 100 x (inhibitor? no enzyme control)/(Me2S0 control? no enzyme control) [4].
Cell Assay
The Wnt/beta-catenin reporter assay was performed with the M50 Super 8x TOPFlash and M51 Super 8x FOPFlash vector containing the firefly luciferase gene under the control of TCF/LEF binding sites or mutated bindings sites. 12,500 cells were seeded overnight on gelatine-coated 96-well plates in LIF-containing ES cell medium. On the next day, the cells were transfected using Lipofectamine with one of the aforementioned vectors plus pGL4.75 [hRluc/CMV] encoding the renilla luciferase reporter gene hRluc as a transfection control. Six hours after transfection the medium was changed to medium devoid of LIF, with reduced serum, and supplemented with 5 uM CHIR-99021. The Dual-Luciferase? reporter assay system.
Kinases were purified from SF9 cells through the use of their His or Glu tag. Glutagged proteins were purified as described, and His-tagged proteins were purified according to the manufacturer's instructions. Kinase assays were performed in 96-well plates with appropriate peptide substrates in a 300-1J1 reaction buffer (variations on 50 mM Tris-HCI, pH 7.5, 10 mM MgCl2, 1 mM EGTA, 1 mM dithiothreitol, 25 mM~-glycerophosphate, 1 mM NaF, and 0.01% bovine serum albumin). Peptides had Km values from 1 to 100 uM. CHIR 99021 or CHIR GSKIA was added in 3.5 ul of Me2SO, followed by ATP to a final concentration of 11JM. After incubation, triplicate 100-u1 aliquots were transferred to Combiplate 8 plates containing 100 ul/well of 50 uM ATP and 20 mM EDTA. After 1 hour, the wells were rinsed five times with phosphate-buffered saline, filled with 200 IJI of scintillation fluid, sealed, and counted in a scintillation counter 30 min later. All of the steps were at room temperature. The percentage of inhibition was calculated as 100 x (inhibitor? no enzyme control)/(Me2S0 control? no enzyme control) [4].
Cell Assay
The Wnt/beta-catenin reporter assay was performed with the M50 Super 8x TOPFlash and M51 Super 8x FOPFlash vector containing the firefly luciferase gene under the control of TCF/LEF binding sites or mutated bindings sites. 12,500 cells were seeded overnight on gelatine-coated 96-well plates in LIF-containing ES cell medium. On the next day, the cells were transfected using Lipofectamine with one of the aforementioned vectors plus pGL4.75 [hRluc/CMV] encoding the renilla luciferase reporter gene hRluc as a transfection control. Six hours after transfection the medium was changed to medium devoid of LIF, with reduced serum, and supplemented with 5 uM CHIR-99021. The Dual-Luciferase? reporter assay system.
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Product Notes
The CHIR-99021 (Catalog #AAA577666) is an Inhibitor and is intended for research purposes only. The product is available for immediate purchase. It is sometimes possible for the material contained within the vial of "CHIR-99021, Inhibitor" to become dispersed throughout the inside of the vial, particularly around the seal of said vial, during shipment and storage. We always suggest centrifuging these vials to consolidate all of the liquid away from the lid and to the bottom of the vial prior to opening. Please be advised that certain products may require dry ice for shipping and that, if this is the case, an additional dry ice fee may also be required.Precautions
All products in the AAA Biotech catalog are strictly for research-use only, and are absolutely not suitable for use in any sort of medical, therapeutic, prophylactic, in-vivo, or diagnostic capacity. By purchasing a product from AAA Biotech, you are explicitly certifying that said products will be properly tested and used in line with industry standard. AAA Biotech and its authorized distribution partners reserve the right to refuse to fulfill any order if we have any indication that a purchaser may be intending to use a product outside of our accepted criteria.Disclaimer
Though we do strive to guarantee the information represented in this datasheet, AAA Biotech cannot be held responsible for any oversights or imprecisions. AAA Biotech reserves the right to adjust any aspect of this datasheet at any time and without notice. It is the responsibility of the customer to inform AAA Biotech of any product performance issues observed or experienced within 30 days of receipt of said product. To see additional details on this or any of our other policies, please see our Terms & Conditions page.Item has been added to Shopping Cart
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