Modular, Antibody-free Time-Resolved LRET Kinase Assay Enabled by Quantum Dots and Tb(3+)-sensitizing Peptides.
Modular, Antibody-free Time-Resolved LRET Kinase Assay Enabled by Quantum Dots and Tb(3+)-sensitizing Peptides.
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DOI：
10.1038/srep28971
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发表时间：
2016-07-18
影响因子：
4.6
通讯作者：
Parker LL
中科院分区：
文献类型：
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作者：
Cui W;Parker LL
荧光药物筛选试验对于发现酪氨酸激酶抑制剂是必不可少的。在这里，我们展示了一种灵活的、无抗体的tr-LRET激酶检测策略，该策略通过链霉亲和素包裹的量子点(QD)受体和生物素化的、Tb3+敏化的多肽供体的组合实现。通过利用Tb3+和Qd的光谱特征以及链霉亲和素-生物素相互作用的高结合亲和力，我们以模块化的方式实现了对激酶活性的多路检测，而不需要对每个肽底物进行额外的共价标记。这一策略与高通量筛选相兼容，并应适应快速变化的工作流程和发现激酶抑制剂所涉及的靶点。
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, Tb3+ sensitizing peptide donors. By exploiting the spectral features of Tb3+ 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.