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However, most XL\u2010MS methods face significant sample loss during enrichment, limiting their applications to limited\u2010quantity samples, and suffer from poor reproducibility (20%\u201340%), hindering precise quantification. To overcome these challenges, we developed a novel membrane\u2010permeable cross\u2010linker, 2,6\u2010dimethylpiperidine disuccinimidyl tridecanoate (DPST), in which the dimethylpiperidinyl group enables one\u2010step enrichment of cross\u2010linked peptides via tandem mass tags (TMTs) antibody approach, eliminating sample loss from multi\u2010step processes and allowing analysis from as few as 1E4 cells. DPST also allows the light and heavy isotopic labeling of cross\u2010linked samples at the cellular level, which reduces inaccuracies from multi\u2010step preparations. This generates reporter ions for precise MS2 quantification, improving the signal\u2010to\u2010noise ratio without increasing spectral complexity. Using DPST, we analyzed cross\u2010links in primary neurons from single fetal mice and quantified the transient and weak interactions in dynamic liquid\u2013liquid phase separation (LLPS) environments. Additionally, DPST's design supports multiple isotopic labeling configurations (e.g., 6\u2010plex, 10\u2010plex). Therefore, DPST provides a scalable and robust tool for in vivo XL\u2010MS\u2010based qualitative and quantitative analysis of living cells, even with limited sample quantities.<\/jats:p>","DOI":"10.1002\/anie.202501845","type":"journal-article","created":{"date-parts":[[2025,5,19]],"date-time":"2025-05-19T10:26:10Z","timestamp":1747650370000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["One\u2010Step Enrichment and Quantitative Analysis of In Vivo Protein Complexes via Dimethylpiperidine Cross\u2010Linker DPST"],"prefix":"10.1002","volume":"64","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3042-2584","authenticated-orcid":false,"given":"Jing","family":"Chen","sequence":"first","affiliation":[{"name":"Department of Chemical Physics School of Chemistry and Materials Science University of Science and Technology of China  96 Jinzhai Road Hefei 230026 China"},{"name":"State Key Laboratory of Medical Proteomics 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