Rapid characterization of N‐linked glycans from secreted and gel‐purified monoclonal antibodies using MALDI‐ToF mass spectrometry

Recombinant monoclonal antibodies (MAbs) are increasingly being used for therapeutic use and correct glycosylation of these MAbs is essential for their correct function. Glycosylation profiles are host cell‐ and antibody class‐dependent and can change over culture time and environmental conditions....

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Bibliographic Details
Published in:Biotechnology and bioengineering 2010-12, Vol.107 (5), p.902-908
Main Authors: Hansen, Rasmus, Dickson, Alan J, Goodacre, Royston, Stephens, Gill M, Sellick, Christopher A
Format: Article
Language:English
Subjects:
Animals
Antibodies
Antibodies, Monoclonal - chemistry
Antibodies, Monoclonal - isolation & purification
Antibodies, Monoclonal - metabolism
antibody
Automation
Biological and medical sciences
Biotechnology
Cell Line
Cells
deglycosylation
Electrophoresis, Polyacrylamide Gel
Fundamental and applied biological sciences. Psychology
Glycan
Glycoproteins
Glycosylation
Health. Pharmaceutical industry
IgG1
IgG4
in-gel analysis
Industrial applications and implications. Economical aspects
MALDI-ToF-MS
Mass spectrometry
Monoclonal antibodies
Polysaccharides - analysis
Production of active biomolecules
Recombinant
Reproduction
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
Workflow
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Summary:Recombinant monoclonal antibodies (MAbs) are increasingly being used for therapeutic use and correct glycosylation of these MAbs is essential for their correct function. Glycosylation profiles are host cell‐ and antibody class‐dependent and can change over culture time and environmental conditions. Therefore, rapid monitoring of glycan addition/status is of great importance for process validity. We describe two workflows of generally applicability for glycan profiling of purified and gel‐purified MAbs produced in NS0 and CHO cells, in which small‐scale antibody purification and buffer exchange is combined with PNGase F glycan cleavage and graphite HyperCarb desalting. MALDI‐ToF mass spectrometry is used for sensitive detection of glycan forms, with the ability to confirm glycan structures by selective ion fragmentation. Both workflows are rapid, technically simple and amenable to automation, and use in multi‐well formats. Biotechnol. Bioeng. 2010;107: 902-908.
ISSN:0006-3592
1097-0290
1097-0290
DOI:10.1002/bit.22879