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Rare Earth Ion Mediated Fluorescence Accumulation on a Single Microbead: An Ultrasensitive Strategy for the Detection of Protein Kinase Activity at the Single‐Cell Level
Author(s) -
Zhang Xiaobo,
Liu Chenghui,
Wang Honghong,
Wang Hui,
Li Zhengping
Publication year - 2015
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.201507580
Subject(s) - microbead (research) , fluorescence , single cell analysis , chemistry , cell , biophysics , nanotechnology , materials science , biochemistry , biology , physics , optics
A single microbead‐based fluorescence imaging (SBFI) strategy that enables detection of protein kinase activity from single cell lysates is reported. We systematically investigated the ability of various rare earth (RE) ions, immobilized on the microbead, for specific capturing of kinase‐induced phosphopeptides, and Dy 3+ was found to be the most prominent one. Through the efficient concentration of kinase‐induced fluorescent phosphopeptides on a Dy 3+ ‐functionalized single microbead, kinase activity can be detected and quantified by reading the fluorescence on the microbead with a confocal fluorescence microscope. Owing to the extremely specific recognition of Dy 3+ towards phosphopeptides and the highly‐concentrated fluorescence accumulation on only one microbead, ultrahigh sensitivity has been achieved for the SBFI strategy which allows direct kinase analysis at the single‐cell level.