Highly Multiplexed Single‐Cell In Situ Protein Analysis with Cleavable Fluorescent Antibodies. Issue 10 (27th January 2017)

• Record Type: Journal Article
• Authors: Mondal, Manas; Liao, Renjie; Xiao, Lu; Eno, Taylor; Guo, Jia
• Is Part Of: Angewandte Chemie international edition. Volume 56:Issue 10(2017)
• Page Start: 2636
• Page End: 2639
• Publication Date: 2017-01-27
• Subjects: fluorescent probes -- immunoassays -- protein expression -- proteomics; Chemistry -- Periodicals; 540
• DOI: 10.1002/anie.201611641
• Languages: English
• ISSNs: 1433-7851

Abstract

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. Abstract : Reiterative staining : Protein targets were stained with antibodies that had been tethered to fluorophores via a chemically cleavable linker. Through continuous cycles of target staining, fluorescence imaging, and fluorophore cleavage, a large number of different protein targets could be detected in individual cells in situ.