Olga Gornik1, Gordan Lauc. 1. Department of Biochemistry and Molecular Biology, Faculty of Pharmacy and Biochemistry, University of Zagreb, A. Kovacića 1, 10000 Zagreb, Croatia.
Abstract
OBJECTIVES: Glycosylation analysis provides many opportunities for diagnostics, but its complexity hampers its routine application. Aiming to alleviate this problem, we developed a simple assay that can measure sialylation of transferrin directly from serum. DESIGN AND METHODS: Transferrin samples with different levels of sialylation were prepared by desialylation. Enzyme-linked-lectin assay (ELLA) and high-performance anion-exchange chromatography (HPAEC) have been used to analyze transferrin sialylation. Periodate oxidation was used to oxidize carbohydrates on antibodies. RESULTS: ELLA was developed for the analysis of serum transferrin sialylation. Antibodies oxidized in situ with periodate have been used to capture transferrin from serum samples. Sialic acid on transferrin has been detected with Sambucus nigra agglutinin (SNA) lectin. Transferrin samples with different sialylation levels prepared by differential desialylation have been used as standards. Accuracy of the method has been confirmed by comparison to HPAEC analysis. CONCLUSIONS: A rapid and simple ELLA that can be routinely used for the analysis of serum transferrin sialylation has been developed.
OBJECTIVES: Glycosylation analysis provides many opportunities for diagnostics, but its complexity hampers its routine application. Aiming to alleviate this problem, we developed a simple assay that can measure sialylation of transferrin directly from serum. DESIGN AND METHODS: Transferrin samples with different levels of sialylation were prepared by desialylation. Enzyme-linked-lectin assay (ELLA) and high-performance anion-exchange chromatography (HPAEC) have been used to analyze transferrin sialylation. Periodate oxidation was used to oxidize carbohydrates on antibodies. RESULTS: ELLA was developed for the analysis of serum transferrin sialylation. Antibodies oxidized in situ with periodate have been used to capture transferrin from serum samples. Sialic acid on transferrin has been detected with Sambucus nigra agglutinin (SNA) lectin. Transferrin samples with different sialylation levels prepared by differential desialylation have been used as standards. Accuracy of the method has been confirmed by comparison to HPAEC analysis. CONCLUSIONS: A rapid and simple ELLA that can be routinely used for the analysis of serum transferrin sialylation has been developed.
Authors: Osmond D Rebello; Richard A Gardner; Paulina A Urbanowicz; David N Bolam; Lucy I Crouch; David Falck; Daniel I R Spencer Journal: Glycoconj J Date: 2020-10-16 Impact factor: 2.916