title: Modular, Antibody-free Time-Resolved LRET Kinase Assay Enabled by Quantum Dots and Tb(3+)-sensitizing Peptides.
abstract: Fluorescent drug screening assays are essential for tyrosine kinase inhibitor discovery. Here we demonstrate a flexible, antibody-free TR-LRET kinase assay strategy that is enabled by the combination of streptavidin-coated quantum dot (QD) acceptors and biotinylated, Tb(3+) sensitizing peptide donors. By exploiting the spectral features of Tb(3+) and QD, and the high binding affinity of the streptavidin-biotin interaction, we achieved multiplexed detection of kinase activity in a modular fashion without requiring additional covalent labeling of each peptide substrate. This strategy is compatible with high-throughput screening, and should be adaptable to the rapidly changing workflows and targets involved in kinase inhibitor discovery.
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 7-194 MCB Building, 420 Washington Ave SE, Minneapolis, MN 55455 USA.|Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, Purdue Center for Cancer Research, Purdue University, West Lafayette, IN 47907 USA.
authors.forename: Wei
authors.initials: W
authors.lastname: Cui
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 7-194 MCB Building, 420 Washington Ave SE, Minneapolis, MN 55455 USA.
authors.forename: Laurie L
authors.initials: LL
authors.lastname: Parker
mesh_terms: D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004735:Energy Transfer; D008163:Luminescent Measurements / Q000379:methods*; D047428:Protein Kinase Inhibitors / Q000302:isolation & purification*; D011505:Protein-Tyrosine Kinases / Q000032:analysis* / Q000037:antagonists & inhibitors*; D045663:Quantum Dots / Q000378:metabolism*
chemical_list: D047428:Protein Kinase Inhibitors; D011505:Protein-Tyrosine Kinases