Cross Validation of Liquid Chromatography-Mass Spectrometry and Lectin Array for Monitoring Glycosylation in Fed-Batch Glycoprotein Production

• Glycosylation analysis of recombinant glycoproteins is of importance for the biopharmaceutical industry and the production of glycoprotein pharmaceuticals. A commercially available lectin array technology was evaluated for its ability to present a reproducible fingerprint of a recombinant CTLY4-IgG fusion glycoprotein expressed in large scale CHO-cell fermentation. The glycosylation prediction from the array was compared to traditional negative mode capillary LC-MS of released oligosaccharides. It was shown that both methods provide data that allow samples to be distinguished by their glycosylation pattern. This included information about sialylation, the presence of reducing terminal galactose beta 1-, terminal N-acetylglucosamine beta 1-, and antennary distribution. With both methods it was found that a general trend of increased sialylation was associated with an increase of the antenna and reduced amount of terminal galactose beta 1-, while N-acetylglucosamine beta 1- was less affected. LC-MS, but not the lectin array, provided valuable information about the sialic acid isoforms present, including N-acetylneuraminic acid, N-glycolylneuraminic acid and their O-acetylated versions. Detected small amounts of high-mannose structures by LC-MS correlated with the detection of the same epitope by the lectin array.

Subject headings

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinsk bioteknologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Medical Biotechnology (hsv//eng)

Keyword

Glycosylation

High mannose

Acetylation

N-linked oligosaccharides

Lectin array

Mass spectrometry

hamster ovary cells

protein

microarray

glycomics

strategy

glycans

oligosaccharides

expression

antibodies

ammonia

Publication and Content Type

ref (subject category)

art (subject category)