Many commonly employed strategies to map kinase activities in live cells require expression of genetically encoded proteins (e.g. FRET sensors). In this work, we describe the development and preliminary application of a set of cell-penetrating, fluorophore labelled peptide substrates for fluorescence lifetime imaging (FLIM) of Abl and Src-family kinase activities. These probes do not rely on FRET pairs or genetically-encoded protein expression. We further demonstrate probe multiplexing and pixel-by-pixel quantification to estimate the relative proportion of modified probe, suggesting that this strategy will be useful for detailed mapping of single cell and subcellular dynamics of multiple kinases concurrently in live cells.
Bibliographical noteFunding Information:
Cells used in this work were a kind gift from Prof. Robert Geahlen (Purdue University Department of Medicinal Chemistry and Molecular Pharmacology). This work was supported by the Purdue Center for Cancer Research (NPD and JI) and the NCI/NIH (R01CA182543 and R33CA217780 to LLP).
© The Royal Society of Chemistry.