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Croatica Chemica Acta, Vol.89 No.2 June 2016. -

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 N-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 N-glycosylation and managed to increase the sensitivity for approximately three times compared to initial experimental conditions, with very good reproducibility.

This work is licensed under a Creative Commons Attribution 4.0 International License.

high-throughput; immunoaffinity chromatography; monoliths; N-glycosylation; oriented antibody immobilization; transferrin



Autor: Irena Trbojević-Akmačić - ; Genos Glycoscience Research Laboratory, Hondlova 2-11, HR-10000 Zagreb, Croatia Blaž Nemec - ; BI

Fuente: http://hrcak.srce.hr/



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