biostudies-literatureUnknown6(11)Vreulz BInteractions between cell surface glycans and lectins mediate vital biological processes, yet their characterization is hindered by the low affinity of these binding events. While photoaffinity labeling can capture these interactions, traditional custom probes often demand tedious synthesis, are limited to simple glycans, and lack versatility. To overcome these limitations, we report a trifunctional scaffold enabling modular assembly of glycan probes. This scaffold integrates orthogonal sites for: (i) efficient late-stage ligation of native oligosaccharides <i>via</i> an <i>N</i>-alkoxy-amine, preserving glycan structure; (ii) flexible amide coupling of various photocrosslinkers, including a recently developed fluorogenic azidocoumarin for traceable labeling; and (iii) conjugation to reporRSC chemical biology1779-1786https://www.ebi.ac.uk/biostudies/studies/S-EPMC12459285biostudies-literatureA trifunctional probe for generation of fluorogenic glycan-photocrosslinker conjugates.PMC12459285Cecioni SVreulz BDe Crozals DfalseA trifunctional probe for generation of fluorogenic glycan-photocrosslinker conjugates.Interactions between cell surface glycans and lectins mediate vital biological processes, yet their characterization is hindered by the low affinity of these binding events. While photoaffinity labeling can capture these interactions, traditional custom probes often demand tedious synthesis, are limited to simple glycans, and lack versatility. To overcome these limitations, we report a trifunctional scaffold enabling modular assembly of glycan probes. This scaffold integrates orthogonal sites for: (i) efficient late-stage ligation of native oligosaccharides <i>via</i> an <i>N</i>-alkoxy-amine, preserving glycan structure; (ii) flexible amide coupling of various photocrosslinkers, including a recently developed fluorogenic azidocoumarin for traceable labeling; and (iii) conjugation to repor2025-01-01T00:00:00Z2025 Oct2026-06-05T08:45:29.453Z2026-05-14T03:12:18.036ZS-EPMC124592854100054210.1039/d5cb00206k