title: Mass spectrometry and HPLC with fluorescent detection-based orthogonal approaches to characterize N-linked oligosaccharides of recombinant monoclonal antibodies.
abstract: A number of HPLC and mass spectrometric techniques are used to characterize post-translational modification in recombinant monoclonal antibodies (MAbs) using the intact glycoprotein and free glycans. LC separation utilizing fluorescent detection technique allows tentative structural assignment of MAb oligosaccharides. Intact molecular weight analysis via electrospray allows for an accurate mass determination and observation of the native glycoform mass envelope. N-linked oligosaccharides are then analyzed by MALDI-ToF. Their structures are further confirmed by analyzing the fragmentation patterns formed by MS/MS. All these techniques provide useful information when performed in isolation. However, the combined information allows for definitive and robust characterization of the N-linked glycans from recombinant MAbs.
authors: Alexion Pharmaceuticals, Cheshire, CT, USA.
authors: Adam W
authors: AW
authors: Lucka
authors: Bruce R
authors: BR
authors: Kilgore
authors: Rekha
authors: R
authors: Patel
authors: Bruce A
authors: BA
authors: Andrien
authors: Shirish T
authors: ST
authors: Dhume
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D009844:Oligosaccharides / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal; D009844:Oligosaccharides; D011994:Recombinant Proteins