inhibitory κB kinase ε
5-20mU of IKKε (diluted in 50mM Tris (pH 7.5), 0.1mM EGTA, 1mg/ml BSA) is assayed against MBP in a final volume of 25.5µl containing 50mM Tris (pH 7.5), 0.1mM EGTA, 0.33mg/ml MBP, 10 mM magnesium acetate and 0.05 mM [33P-γ-ATP]( 500-1000 cpm/pmole) and incubated for 30 mins at room temperature. Assays are stopped by addition of 5µl of 0.5M (3%) orthophosphoric acid. Assays are harvested onto P81 Unifilter plates using a wash buffer of 50mM orthophosphoric acid.
GST-IKK_epsilon.pdf (88.41 KB)