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. 1992 Aug 1;285(Pt 3):839–845. doi: 10.1042/bj2850839

Different culture methods lead to differences in glycosylation of a murine IgG monoclonal antibody.

T P Patel 1, R B Parekh 1, B J Moellering 1, C P Prior 1
PMCID: PMC1132872  PMID: 1497622

Abstract

A monoclonal IgG-1 was produced by culture of a murine hybridoma (3.8.6) by three different methods, namely culture in ascites, in serum-free media and in serum-supplemented media. IgG-1 was purified to homogeneity (as judged by SDS/PAGE under reducing conditions) from each medium by ion-exchange chromatography and h.p.l.c. Protein A chromatography. Oligosaccharides were released from each IgG-1 preparation by hydrazinolysis and radiolabelled by reduction with alkaline sodium borotritide, and 'profile' analysis of the radiolabelled oligosaccharide alditols was performed by a combination of paper electrophoresis and gel-filtration chromatography. This analysis indicated clear and reproducible differences in the glycosylation patterns of the three IgG-1 preparations. Sequential exoglycosidase analysis of individual oligosaccharides derived from each IgG-1 preparation was used to define these differences. Ascites-derived material differed from serum-free-culture-derived material only with respect to the content of sialic acid. IgG-1 derived from culture in serum-containing media had an intermediate sialic acid content and a lower incidence of outer-arm galactosylation than the other two preparations. These differences in glycosylation could not be induced in any IgG-1 preparation by incubating purified IgG-1 with ascites or culture medium. It is concluded that the glycosylation pattern of a secreted monoclonal IgG is dependent on the culture method employed to obtain it.

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Selected References

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