{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T17:16:30Z","timestamp":1788455790375,"version":"build-2803163510"},"reference-count":40,"publisher":"American Chemical Society (ACS)","issue":"9","license":[{"start":{"date-parts":[[2026,7,6]],"date-time":"2026-07-06T00:00:00Z","timestamp":1783296000000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-029"},{"start":{"date-parts":[[2026,7,6]],"date-time":"2026-07-06T00:00:00Z","timestamp":1783296000000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-037"},{"start":{"date-parts":[[2026,7,6]],"date-time":"2026-07-06T00:00:00Z","timestamp":1783296000000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-045"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,9,2]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Glycosylation of monoclonal antibodies is a key determinant of their efficacy, stability, and safety, requiring analytical methods capable of resolving site-specific heterogeneity with high sensitivity. In this work, we present a Protein A-assisted, domain-resolved glycan profiling approach for the mass spectrometry-based structural elucidation and quantification of N-glycans linked to Fc and Fab in cetuximab. Antibody domains were selectively extracted using Protein A affinity separation, allowing for independent enzymatic release of N-glycans from Fc and Fab sections, followed by high-resolution mass spectrometry for detailed glycan profiling and comparative characterization of Fc- and Fab-associated glycan populations. In comparison to conventional total glycan analysis, our technique decreases spectral complexity and allows for obvious distinction of domain-specific glycosylation patterns. Crucially, domain-specific released glycan profiling improves the detection and precise measurement of low-abundance glycoforms, such as sialylated, high-mannose, and afucosylated species that frequently remain hidden in conventional approaches. The proposed approach facilitates domain-specific characterization of glycan distributions across antibody regions by reducing analytical complexity and improving interpretation of Fc- and Fab-associated glycosylation patterns. Our results show that a reliable and repeatable platform for thorough, domain-level glycan characterization and quantification is available, supporting applications in quality control and biosimilar comparability studies, and in general during biopharmaceutical development.<\/jats:p>","DOI":"10.1021\/jasms.6c00136","type":"journal-article","created":{"date-parts":[[2026,7,6]],"date-time":"2026-07-06T15:54:23Z","timestamp":1783353263000},"page":"2094-2102","source":"Crossref","is-referenced-by-count":0,"title":["Structural Elucidation\nof Fc- and Fab-Associated\n                    <i>N<\/i>\n                    -Glycans in Cetuximab\nUsing Protein A-Assisted\nDomain-Resolved Glycan Profiling Using Mass Spectrometry"],"prefix":"10.1021","volume":"37","author":[{"given":"Divya","family":"Chauhan","sequence":"first","affiliation":[{"name":"Indian Institute of Technology , , Hauz Khas, , ,","place":["New Delhi, Delhi, India, 110016"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tarun","family":"Shivach","sequence":"additional","affiliation":[{"name":"Indian Institute of Technology , , Hauz Khas, , ,","place":["New Delhi, Delhi, India, 110016"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5913-4244","authenticated-orcid":true,"given":"Anurag S.","family":"Rathore","sequence":"additional","affiliation":[{"name":"Indian Institute of Technology , , Hauz Khas, , ,","place":["New Delhi, Delhi, India, 110016"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,7,6]]},"reference":[{"key":"2026090311504955000_cit1","doi-asserted-by":"publisher","first-page":"54","DOI":"10.1208\/s12248-026-01207-0","article-title":"Glycan Profiles of\nFDA - Approved Therapeutic Antibodies: Insights from Regulatory Submissions","volume":"28","author":"Luo","year":"2026","journal-title":"AAPS J."},{"key":"2026090311504955000_cit2","doi-asserted-by":"publisher","first-page":"7320","DOI":"10.3390\/app14167320","article-title":"Comparison\nof Four Rapid N-Glycan Analytical Methods\nand Great Application Potential in Cell Line Development","volume":"14","author":"Jin","year":"2024","journal-title":"Appl. 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Chem."},{"key":"2026090311504955000_cit7","doi-asserted-by":"publisher","first-page":"5393","DOI":"10.3390\/molecules29225393","article-title":"LC-MS\/MS-Based\nSite-Specific N-Glycosylation Analysis of VEGFR-IgG Fusion Protein\nfor Sialylation Assessment Across IEF Fractions","volume":"29","author":"Kim","year":"2024","journal-title":"Molecules"},{"key":"2026090311504955000_cit8","first-page":"1","volume-title":"High-Throughput Glycomics\nand Glycoproteomics: Methods and Protocols","author":"Gordan","year":"2018"},{"key":"2026090311504955000_cit9","doi-asserted-by":"publisher","first-page":"4859","DOI":"10.1021\/pr500460k","article-title":"LC \u2013 MS Pro\nFi Ling of N - Glycans Derived from Human Serum Samples for Biomarker\nDiscovery in Hepatocellular Carcinoma","volume":"13","author":"Tsai","year":"2014","journal-title":"J. Proteome\nRes."},{"key":"2026090311504955000_cit10","unstructured":"Yang, H.; Warren, W. J.; Koza, S. 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