Chromatographic monoliths for high-throughput immunoaffinity isolation of transferrin from human plasma

Scientific paper - Original scientific paper

• Author: Irena Trbojević-Akmačić
• Author: Blaž Nemec
• Author: Urška Vidic
• Author: Suzana Malić
• Author: Karmela Miklić
• Author: Urh Černigoj
• Author: Jana Vidič
• Author: Nika Lendero Krajnc
• Author: Aleš Štrancar
• Author: Gordan Lauc
• Author: Tihana Lenac Roviš
• Author: Maja Pučić-Baković https://orcid.org/0000-0003-0866-623X
• Author's institution: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of biochemistry and molecular biology)
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Medical Biochemistry

Abstract (english)

Changes in protein glycosylation are related to different diseases and have a potential as diagnostic and prognostic disease biomarkers. Transferrin (Tf) glycosylation changes are common marker for congenital disorders of glycosylation. However, biological interindividual variability of Tf <i>N</i>-glycosylation and genes involved in glycosylation regulation are not known. Therefore, high-throughput Tf isolation method and large scale glycosylation studies are needed in order to address these questions. Due to their unique chromatographic properties, the use of chromatographic monoliths enables very fast analysis cycle, thus significantly increasing sample preparation throughput. Here, we are describing characterization of novel immunoaffinity-based monolithic columns in a 96-well plate format for specific high-throughput purification of human Tf from blood plasma. We optimized the isolation and glycan preparation procedure for subsequent ultra performance liquid chromatography (UPLC) analysis of Tf <i>N</i>-glycosylation and managed to increase the sensitivity for approximately three times compared to initial experimental conditions, with very good reproducibility.
<br><a rel="license" href="http://creativecommons.org/licenses/by/4.0/"><img alt="Creative Commons License" style="border-width:0" src="https://i.creativecommons.org/l/by/4.0/80x15.png" /></a> This work is licensed under a <a rel="license" href="http://creativecommons.org/licenses/by/4.0/">Creative Commons Attribution 4.0 International License</a>.

• Keywords (english): high-throughput; immunoaffinity chromatography; monoliths; N-glycosylation; oriented antibody immobilization; transferrin
• Language: english
• Publication type: Scientific paper - Original scientific paper
• Publication status: Published
• Peer review: Peer review - international
• Publication version: Published version
• Journal title: Croatica Chemica Acta
• Numbering: vol. 89, no. 2, pp. 203-211
• p-ISSN: 0011-1643
• e-ISSN: 1334-417X
• DOI: https://doi.org/10.5562/cca2815
• URN:NBN: urn:nbn:hr:163:679166
• Publication: 2016
• Document URL: https://hrcak.srce.hr/166006
• Type of resource: Text
• Access conditions: Open access
• Created on: 2020-04-17 13:38:40