{"doi":"10.1101/2025.10.16.682811","title":"Time-Resolved Tracking and Secretome Mapping of Intracellular, Cell Surface, and Extracellular Glycoconjugates with a Mini-Tetrazine N-Acetylglucosamine","abstract":"Glycosylation is instrumental in governing essential biological processes that dictate health outcomes. Herein we describe a mini-tetrazine N-acetylglucosamine (HTz-GlcNAc) that is incorporated into N-linked glycans and through rapid live-cell labeling with trans-cyclooctene (TCO) reagents enables real-time visualization and evaluation of glycoconjugate trafficking in living cells. We utilize cell-permeable and impermeable TCO-fluorophores to perform time-resolved, multicolor live labeling of cell-surface glycoconjugates, as well as intracellular, and internalized glycoconjugates. We observe intracellular glycoconjugate trafficking throughout the secretory pathway of live cells in real-time, as well as cell surface glycan turnover and extracellular vesicle secretion across three days. In the toolkit of probes for metabolic oligosaccharide engineering, HTz-GlcNAc brings the capability of precise interrogation of intracellular and extracellular glycoconjugate trafficking, and selective tagging of intra-and-extracellular vesicles, while providing a handle for the purification and enrichment of glycans for immunoblot and proteomic analyses.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":578201,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9587,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":676877,"name":"Mary W. N. Burns","orcid":"0000-0003-0712-237X","position":1,"is_corresponding":false},{"id":1488250,"name":"Sarah L. Janney","orcid":null,"position":2,"is_corresponding":false},{"id":1488251,"name":"Ashley M. Stark","orcid":null,"position":3,"is_corresponding":false},{"id":1487843,"name":"Feng Cai","orcid":"0000-0003-4473-6477","position":4,"is_corresponding":false},{"id":1039170,"name":"Timothy Chaya","orcid":"0000-0003-4821-0317","position":5,"is_corresponding":false},{"id":308356,"name":"Catherine L. Grimes","orcid":"0000-0002-0586-2879","position":6,"is_corresponding":false},{"id":268923,"name":"Jennifer J. Kohler","orcid":"0000-0001-5373-3329","position":7,"is_corresponding":false},{"id":324007,"name":"Joseph M. Fox","orcid":"0000-0002-8258-1640","position":8,"is_corresponding":false},{"id":938745,"name":"Ashlyn S. Hillman","orcid":null,"position":0,"is_corresponding":true}],"reference_count":0,"raw_metadata":null,"created_at":"2026-07-19T02:58:20.638044Z","pmid":"41279695","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}