Fig 1.
Expression and polymer distribution of IgM antibodies produced in CHO DG44 and HEK293E cells.
(A) IgM concentrations are shown as bars and viable cell concentrations are represented as dots. Stable expression in CHO cells was performed in triplicates. (B) Silver staining under non-reducing conditions of purified IgM012, IgM012_GL and IgM617 produced stably in CHO cells and transiently in HEK cells. Pentameric and dimeric forms are indicated with arrows.
Table 1.
Comparison of site-specific glycosylation profile at GS1-3 of IgM produced stably in CHO cells and transiently in HEK cells.
Relative abundance [%] of N-glycan types is shown.
Table 2.
Comparison of oligomannose structures at GS4 of IgM produced stably in CHO cells and transiently in HEK cells.
Relative abundance [%] of N-glycan structures is shown.
Fig 2.
Non-processed images of negative stain transmission electron microscopy images of IgM antibody.
Representative fields of particles with a 100 nm scale bar in the upper pictures. Magnified views of some individual molecules are shown in the lower panels for all three model antibodies produced in HEK293E cells: (A) IgM012, (B) IgM012_GL, (C) IgM617.
Fig 3.
Complement activation via IgM and C1q interaction.
Indicated IgMs were coated on the plate and incubated with normal human serum (NHS), NHS depleted of C1q (NHSΔ) or NHSΔ reconstituted with C1q. Polyclonal IgM (pIgM, black) as well as IgM012 (blue), IgM012_GL (green) and IgM617 (red) produced stably in CHO DG44 (darker colors) or transiently in HEK293E cells (lighter colors). Serum C4 cleavage by the IgM-bound activated C1 complex results in C4b deposition detected by a C4b specific antibody. All samples were analysed in duplicates.