Profiling N-linked oligosaccharides from IgG by high-performance anion-exchange chromatography with pulsed amperometric detection

Glycobiology
Jeffrey S RohrerDeanna C Hurum

Abstract

Understanding and characterizing protein therapeutic glycosylation is important with growing evidence that glycosylation impacts biological efficacy, pharmacokinetics and cellular toxicity. Protein expression systems and reactor conditions can impact glycosylation, leading to potentially undesirable glycosylation. For example, high-mannose species may be present, which are atypical of human antibody glycosylation. Their presence in the Fc domain has been linked to increased serum clearance of immunoglobulin G (IgG) antibodies. High-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) is an effective tool for determining glycans present in glycoprotein therapeutics. We report an improved HPAE-PAD method for IgG oligosaccharide separation. The neutral glycans are well resolved, including separation of high-mannose species from typical human IgG glycans. Oligosaccharide identification was performed by comparison to known standards in conjunction with selective exoglycosidase digestion of both standards and released glycans. Retention times (RTs) of known glycans were compared with the retention times of maltose, maltotriose and maltotetraose standards to define a retention index value for each gl...Continue Reading

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Citations

Apr 4, 2017·Electrophoresis·Lucas VeillonYehia Mechref
Aug 22, 2017·Analytical Methods : Advancing Methods and Applications·Gabe NagyNicola L B Pohl
Jul 29, 2020·Frontiers in Chemistry·Qing ZhangXuezheng Song
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Jan 21, 2017·Analytical Chemistry·Stefan GaunitzMilos V Novotny

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