Protocols assessing kinase activity generally include radioactive methods, fluorescent polarization and the use of anti-phosphorylation antibodies. The present study described the application of a luminescence-based ATP quantitative reagent, Kinase-Glo, in evaluating the activity of cyclin-dependent kinase 2 (CDK2). Different amounts of immunoprecipitated CDK2 retrieved from HeLa extracts and different concentrations of the CDK2 inhibitor, roscovitine, were applied to test the sensitivity of Kinase-Glo reagent. The results demonstrated that Kinase-Glo reagent was sensitive to detect the alterations of CDK2 activity and suggested it to be applied in assessing kinase activity and for inhibitor optimization in vitro.
Key words: Cyclin-dependent kinase 2 (CDK2), kinase assay, Kinase-Glo, relative light unit (RLU), luminescence.
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