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Valegh Faid and colleagues at LFB Biotechnologies in France have developed and published an assay to study antibody disulphide bonds using middle-up LC-MS (Faid et al. 2017). In the paper, the combination of FabRICATOR® for digestion below the hinge and FabALACTICA™ for digestion above the hinge, generated three fragments from a human IgG1 antibody; the hinge…
"GingisKHAN™ protease cleavage allows a high-throughput antibody to Fab conversion enabling direct functional assessment during lead identification of human monoclonal and bispecific IgG1 antibodies." In this paper, the scientists at Roche Innovation Center Munich, Moelleken et al, discuss the use of GingisKHAN in antibody lead selection screening and affinity measurements. The binding strength of…
In a collaboration between Universite de Strasbourg, Pierre-Fabre, and Catalent Biologics a multilevel mass spectrometry analysis of a site-specific conjugated ADC was recently published in the journal mAbs. As a first step in the analysis, FabRICATOR (IdeS) was used to perform middle-up analysis and confirm the drug to antibody ratio, DAR. The scientists write on p.7:…
A team of scientist from Quality Assistance in collaboration with Alain Beck from Pierre Fabre, have published a detailed article highlighting the analytical strategies for both N- and O-linked glycan analysis of biotherapeutics using LC/MS. The workflows for glycan analysis on antibodies includes FabRICATOR digestion and study of the Fc/2 fragment for identification of the Fc glycoforms.…
Biosimilars are gaining in popularity as patents of innovator drugs are expiring. The analytical strategies to characterize and assess the similarity between the innovator product and the biosimilar often involve liquid chromatography and mass spectrometry (LC/MS). In a recent paper from the Freie Universität Berlin, Montacir et al. studied Rituximab and follow-on molecules using FabRICATOR digestion, reduction, and…
Oxidation of methionine residues in the Fc region of a therapeutic antibody may affect the binding of the antibody to Protein A and FcRn leading to difficulties in purification or increased clearence in vivo. For the variable regions of the antibody, oxidation may affect antigen binding or lead to increased immunogenicity. For these reasons, the…
Antibodies are important tools in several scientific research areas, and sometimes it is necessary to cleave antibodies to generate antigen binding (Fab) fragments. The Fab fragments can be used within e.g. imaging, removal of effector functions, infection biology, binding studies, and mass spectrometry. In a recently published book (“Bacterial Pathogenesis, Methods and Protocols”, Humana Press,…
Researchers at LFB Biotechnologies have published the first paper using the Genovis SmartEnzyme GingisKHAN. GingisKHAN digests human IgG1 antibodies in the upper hinge, resulting in intact Fab and Fc fragments. The paper published in Journal of Chromatography describes how charge variants of therapeutic antibodies are characterized in a long term stability study using ion exchange chromatography coupled to…
The level of core fucosylation on therapeutic antibodies have major impact on antibody dependent cell cytotoxicity. To study the core fucosylation, Suli Liu and Li Zang at Biogen have published a method using deglycosylation by GlycINATOR, reduction of the antibody and LC-MS analysis. Using this methodology the scientist could quantify the fucosylation within 1 h, including sample…
The Genovis team has together with Alain Beck, Senior Director Antibody Physico-Chemistry at Pierre Fabre, published a review article summarizing all the current publications using FabRICATOR (IdeS) for characterization of therapeutic antibodies and related products. "The specificity and robustness of the IdeS digestion is critical for generating a homogenous pool of fragments. Combined with…
Researchers at the University of Geneva led by Davy Guillarme has used a novel form of HPLC column, based on wide-pore hybrid silica bonded with amide ligand, and developed an HILIC analytical workflow for characterization of biopharmaceuticals. The scientists used FabRICATOR to fragment trastuzumab to subunits and studied the fragments using the new column with HILIC separation.…
New references are out using Genovis enzymes to study antibody glycation, pairing of high-mannose glycans and ADC characterization using CE-MS.