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Incorporating Mass Spectrometry into Biologic QC Control Environments for O-Glycan Analysis

Mark Hilliard  Ciara A.McManus  William Alley  Ying Qing Yu  John Gebler  Pauline M Rudd  
【摘要】:O-linked glycans are typically more challenging to analyze than N-linked glycans,partly because of the diversity of the core structures as well as the lack of a universal O-glycanase.Further,most O-glycan workflows are performed on high-end mass spectrometry instruments which are not available in most QC environments.These MS instruments do not provide the quantitative data required by the regulators and there can be a lack of skilled personnel to operate them.With the emergence of O-glycosylated biotherapeutics,it is now necessary to develop alternative workflows;first a more research focused LC-QTof approach and secondly,a more QC compatible methodology employing a novel mass detector,the QDa.Researchers typically employ chemical techniques for O-linked glycan release.Here we present a microwave-assisted O-linked glycan release workflow that can be completed within 12-70 minutes in comparison to traditional beta-elimination methods which takes around 16 hours.The addition of the QDa mass detector allowed for the addition of mass data that is complementary to more traditional optical detectors that are found within the QC environment.Further,we can increase MS sensitivity with the addition of an MS compatible fluorophore which also facilitates the collection of qualitative and quantitative LC-MS data which is not generally represented in current assays.For more complex samples or where accurate mass and MS/MS data is required,our LC-MS method can be transferred effortlessly to more research orientated instruments such as the QTof-MS.
【作者单位】:NIBRT,Foster Avenue,Mount Merrion,Blackrock,Co Dublin Waters Corporation

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