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Assay Method Information |
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| | KinaseGlo Plus Luminescent Assay |
| Description: | A stock solution of 5 % DMSO (BioReagent for molecular biology, Sigma Aldrich) in water was prepared. The CK1 substrate peptide RRKDLHDDEEDEAMSITA (Jena Bioscience, cat. no. PE-206, 2 mM stock solution in water) was diluted in water to a final concentration of 250 μM. CK1δ or CK1Ɛ kinases were diluted with CK1 dilution buffer to 50 ng / µL. In addition, fresh Reaction buffer was prepared containing 250 mM Tris-HCl (pH 7.5), 50 mM MgCl2, 5 mM DTT and 0.5 mg / mL bovine serum albumin (BSA). Compounds were serially diluted in 5 % DMSO solution to the desired concentrations (maximal concentration employed in the assay was 50 µM). Additional samples included blank sample containing Reaction buffer, water and CK1δ / CK1Ɛ kinase and control sample containing Reaction buffer, CK1 substrate peptide and CK1δ / CK1Ɛ kinase. CK1 substrate peptide, Reaction buffer and CK1δ / CK1Ɛ kinase stock solutions were mixed in 1:1:1 ratio. 6 μL of this mixtur |
| Affinity data for this assay | |
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