Relative quantitation of neutral and sialylated N-glycans using stable isotopic labeled d0/d5-benzoyl chloride by...

Quantitative analysis of glycans is an emerging field in glycomic research. Herein we present a rapid and effective dual-labeling strategy, in the combination of isotopic derivatization of N-glycosylamine-based glycans by d0/d5-benzoyl chloride and methylamidation of sialic acids, to relatively quantify both neutral and sialylated N-glycans simultaneously by MALDI-MS. The derivatization efficiencies were increased by microwave-accelerated deglycosylation which not only largely reduce the time of glycoprotein deglycosylation but also inhibit the hydrolysis of intermediate glycosylamines produced by PNGase F digestion. Three model glycoproteins, including RNase B, bovine fetuin and IgG from human serum, were applied to validate this technique. Results showed that the glycans from microgram level of glycoprotein can be successfully quantified with high reproducibility and the whole time of analytical procedure was shortened to 4 h. Furthermore, this proposed method was applied for the comparative analysis of N-glycans from serum of healthy donors and multiple myeloma patients. It was found that five N-glycans may be as the potential biomarkers for rapid detection of early multiple myeloma, indicating the feasibility of this strategy in monitoring subtle quantitative differences of serum glycomics.