GlcNAcylation mapping of single living cells by quantitative SERS imaging

O -GlcNAcylation is involved in many biological processes including cancerization. Nevertheless, its in situ quantification in single living cells is still a bottleneck. Here we develop a quantitative SERS imaging strategy for mapping the O -GlcNAcylation distribution of single living cells. O -GlcNAcylated compounds (OGCs) can be quantified through their in situ azide labeling and then a click reaction competing with azide and Raman reporter labeled 15 nm-gold nanoparticles (AuNPs) for linking to dibenzocyclooctyne labeled 40 nm-AuNPs to produce OGC-negatively correlated SERS signals. The calibration curve obtained in vitro can be conveniently used for detecting OGCs in different areas of single living cells due to the negligible effect of cell medium on the click linkage and Raman signal. This method has been successfully applied in mapping O -GlcNAcylation distribution in different cell lines and monitoring O -GlcNAcylation variation during cell cycling, which demonstrate its great practicability and expansibility in glycosylation related analysis. A quantitative SERS imaging strategy is developed for O -GlcNAcylation mapping of single living cells through a competitive click reaction.