Kinases play a central role in cell signaling due to their protein phosphorylating function. While enzymes of the protein kinase C (PKC) family phosphorylate serine and threonine residues, tyrosine kinases exclusively phosphorylate tyrosines. Their importance in cellular processes such as proliferation and in diseases like cancer or diabetes makes kinases a highly interesting object of research.
This application note presents the reliable detection of activity of either kinase family by the LanthaScreen® TR-FRET assay. It uses kinase-specific substrate peptides labelled with fluorescein. If the peptide is phosphorylated by the specific kinase, a phospho-specific and terbium-coupled antibody can bind. The resulting phosphorylation-dependent proximity of terbium and fluorescein allows for energy transfer. The PHERAstar® simultaneously detects emission of terbium and fluorescein and provides data for a ratiometric analysis. The assay reflected different concentrations of kinases of either PKC or tyrosine type and proves a rapid and reliable assay for kinase