Fig 1.
a) A representative schematic structure of monoclonal antibody and N-glycosylation sites on it. The main glycan moieties of the Fab and Fc fragment were shown in the frame. Structures and the monosaccharides are depicted following the CFG notation; b) flowchart of our method in this study.
Fig 2.
MALDI-TOF MS spectrum of N-glycans enzymatically released from the biosimilar of cetuximab and cetuximab.
a) native N-glycans before mild alkali treatment (pH 10 ammonium hydroxide); b) native N-glycans of the biosimilar after mild alkali treatment; c) native N-glycans from the cetuximab. The cartoons of possible structures of glycans were adapted from Glycoworkbench and structure is depicted following the CFG notation.
Fig 3.
Typical NP-HPLC spectrum of 2-AA labeled glycans from the biosimilar of cetuximab.
a) 2-AA labeled mAbs glycans before mild alkali treatment; b) 2-AA labeled mAbs glycans after mild alkali treatment.
Fig 4.
NanoLC-ESI-MS/MS spectrum of native glycans.
a) MS/MS spectra of m/z 2060 with chemical composition of GlcNAc4Man3Gal2NeuAcLac1; b) MS/MS spectra of m/z 2078 with chemical composition of GlcNAc4Man3Gal2NeuAc1.
Fig 5.
Typical NP-HPLC spectrum of 2-AA labeled glycans from the Fab and Fc fragment of the biosimilar of cetuximab.
a) N-glycans on the Fab fragment; b) N-glycans on the Fc fragment. The compositions and structural schemes of glycans in each chromatographic peak are shown in S2 Table of the Electronic Supplementary Material.