We analysed the oligosaccharides of a human IgM produced by a human-hu
man-mouse hybridoma at each of its five conserved heavy chain glycosyl
ation sites, Consistent with previous reports, this IgM possesses sial
ylated oligosaccharides at Asn171, Asn332 and Asn395, and high-mannose
-type oligosaccharides at Asn402. In contrast to previous reports for
human IgMs, we find that Asn563 is not occupied by oligosaccharide on
perhaps 25% of IgM heavy chains, while occupied Asn563 sites contain b
oth high-mannose-type and sialylated oligosaccharides. These latter re
sults are consistent with the glycosylation at Asn563 previously repor
ted for the mouse MOPC 104E IgM, We demonstrate that both the human hy
bridoma IgM and the mouse MOPC 104E IgM are mixtures of pentamers and
hexamers, raising the possibility that the unique findings concerning
the glycosylation at Asn563 in this study and the previous study of th
e MOPC 104E IgM could be related, at least in part, to the different p
acking requirements of the hexameric geometry and the accessibility of
oligosaccharides in the hexameric geometry for processing to complex
type, In addition, we used high-pH anion-exchange (HPAE) chromatograph
y, neutral anion-exchange chromatography, fluorophore-assisted carbohy
drate electrophoresis and Western blots to compare the oligosaccharide
compositions of the human hybridoma IgM, pooled human serum IgM and t
wo mouse monoclonal IgMs (MOPC 104E and TEPC 183), Of note is the pres
ence of N-glycolylneuraminic acid (NeuGc) and N-acetylneuraminic acid
(NeuAc) at a 2:1 ratio in the oligosaccharides of the human hybridoma
IgM. The presence of both NeuGc and NeuAc complicates the interpretati
on of HPAE chromatographs.