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Highly Multiplexed Single‐Cell In Situ Protein Analysis with Cleavable Fluorescent Antibodies
Author(s) -
Mondal Manas,
Liao Renjie,
Xiao Lu,
Eno Taylor,
Guo Jia
Publication year - 2017
Publication title -
angewandte chemie
Language(s) - English
Resource type - Journals
eISSN - 1521-3757
pISSN - 0044-8249
DOI - 10.1002/ange.201611641
Subject(s) - in situ , fluorophore , fluorescence , single cell analysis , chemistry , linker , green fluorescent protein , cell , biophysics , microbiology and biotechnology , biology , biochemistry , computer science , physics , organic chemistry , quantum mechanics , gene , operating system
Abstract Limitations on the number of proteins that can be quantified in single cells in situ impede advances in our deep understanding of normal cell physiology and disease pathogenesis. Herein, we present a highly multiplexed single‐cell in situ protein analysis approach that is based on chemically cleavable fluorescent antibodies. In this method, antibodies tethered to fluorophores through a novel azide‐based cleavable linker are utilized to detect their protein targets. After fluorescence imaging and data storage, the fluorophores coupled to the antibodies are efficiently cleaved without loss of protein target antigenicity. Upon continuous cycles of target recognition, fluorescence imaging, and fluorophore cleavage, this approach has the potential to quantify over 100 different proteins in individual cells at optical resolution. This single‐cell in situ protein profiling technology will have wide applications in signaling network analysis, molecular diagnosis, and cellular targeted therapies.