Glycan atlassing enables functional tracing of cell state.

The glycocalyx is a complex layer of glycosylated molecules that surrounds all cells in the human body. It is involved in regulating critical cellular processes, including immune response modulation, cell adhesion and host-pathogen interactions. Despite these insights, the functional relationship between the glycocalyx architecture and cellular state has remained elusive, largely due to the structural diversity of glycocalyx constituents and their nanoscale organization. Here we show that DNA-tagged lectin labelling and metabolic oligosaccharide engineering enable multiplexed super-resolution microscopy of the glycocalyx constituents, yielding an atlas of glycocalyx architecture with nanometre resolution. Quantitative analysis of the obtained nanoscale map of glycocalyx constituents facilitates the extraction of characteristic spatial relationships that accurately report on the cellular state. We demonstrate the capacity of our approach, which we term glycan atlassing, across cell and tissue types, ranging from cultured cell lines to primary immune cells, neurons and primary patient tissue. Glycan atlassing establishes a transformative strategy for investigating glycocalyx remodelling in development and disease, potentially enabling the development of glycocalyx-centred targets in diagnosis and therapy.