Sialic Acid Derivatization for the Rapid Subclass- and Sialic Acid Linkage-Specific MALDI-TOF-MS Analysis of IgG Fc-Glycopeptides

Abstract

Matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS) is a highly suitable method for the rapid analysis of IgG glycopeptides, providing a wealth of structural information. A limitation of this approach is that it generates a bias when analyzing sialylated species due to the labile nature of sialic acid glycosidic linkages. One way to overcome this problem is by chemical derivatization of the sialic acids. The method presented here results in both the stabilization of the sialic acids, as well as the differentiation of α2,3- and α2,6-linked sialic acids by mass. Described in this chapter are the isolation of IgG from plasma or serum, tryptic digestion of the samples, derivatization, and finally MALDI-TOF-MS measurement and data analysis.

Keywords: Biopharmaceuticals; Clinical cohorts; Cotton hydrophilic interaction liquid chromatography (HILIC)Matrix assisted laser desorption/ionization (MALDI); Fragment crystallizable (Fc)-glycosylation; Glycopeptide; Immunoglobulin G (IgG); Linkage-specific; Mass spectrometry (MS); N-acetylneuraminic acid; N-glycolylneuraminic acid; Sialic acid; Site-specific; Solid-phase extraction (SPE); Stabilization; Subclass-specific; Time-of-flight (TOF).