Figure 1
Generation of glyco-engineered human IgG1. (a) Schematic representation of glyco-engineered IgG1 antibodies generated in this study with their biantennary glycan on asparagine (Asn) 297 (white star) in the Fc CH2 domain and, for the Fab-glycoform, on Asn 29 and Asn 86 (orange stars) in the VH and VL domain, respectively. (b) Bar graph summarizing Fc-glycoform N-glycan profile of variable glycan end groups (fucose (+/−F), in red; bisecting GlcNAc (+/−B) in cyan; mono-/di-galactose in pale/dark yellow; mono-/di-sialic acid in pale/dark purple) determined by mass spectrometry as percentages (%) of total glycosylation. N-glycan structures are schematically represented; galactose (yellow circle); mannose (green circle); N-acetylglucosamine (blue square); bisecting N-acetylglucosamine (cyan square); fucose (red triangle); sialic acid (purple diamond). (c) Bar graph showing the level of sialic acid of Fab-glycosylated anti-TNP as determined by Sambucus nigra agglutinin (SNA)-lectin ELISA with high sialic acid Fab-glycoform (grey bar) to asialylated (unmodified) IgG1 without a Fab-glycan (white bar). Data are representative of three independent experiments (in triplicate) showing mean ± standard error of the mean (s.e.m.). Statistical analysis was performed by a Two-tailed paired t-test (**P < 0.01).
