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Super resolution microscopy techniques such as stimulated emission depletion microscopy (STED) improves imaging resolution compared to conventional light microscopy. In STED microscopy, super resolution is achieved using photoactivatable dyes that are excited and de-excited selectively with a laser to restrict fluorescense to a specific focal point. While such super resolution methods in combination with immunostaining…
Charge heterogeneity of monoclonal antibodies is an important critical quality attribute that requires close monitoring due to its potential impact on antibody efficacy and immunogenicity. Since the heterogeneity is mostly caused by post translational modifications such as C-terminal lysine clipping, deamidation, glycation, sialic acid or adduct formation, these modifications can pose significant challenges to the…
Genovis launches new GlyCLICK kits for site-specific generation of custom ADCs carrying unique 2-step cleavable linker-payloads from Glykos Finland. Antibody-drug conjugates (ADCs) comprise a new generation of antibody-based biologics that carry drug payloads directly into target cells, allowing for a broadened therapeutic window. The drawbacks of conventional antibody conjugation strategies are rapidly being surpassed by…
Antibodies formulated as solid-state depots can potentially be used for local treatments and minimize the need for large systemic doses. Bevacizumab may for instance be administered locally to control post-operative scarring following glaucoma filtration surgery. A solid-state form would however be required in order to obtain a proper slow release of the antibody.
The formulation of antibodies in mixtures has revealed significant clinical advantages but causes increased analytical challenges. Long-term studies of formulated antibody mixtures over time are both difficult and time consuming. An example of a post-translational modification that could occur during storage is the oxidation of tyrosine that may induce conformational changes of an antibody.
Two complementary determining regions (CDR3) are considered major determinants of antigen-binding specificity that give rise to the human immunoglobulin repertoire with billions of unique antibodies. For de novo sequencing of the human repertoire, circulating antibodies can be analyzed by mass spectrometry after proteolytic cleavage. Complex mixtures such as plasma derived samples are however challenging to…
In contrast to small generic molecules, therapuetic monoclonal antibodies (mAbs) exhibit inherent heterogeneity that may arise during production and formulation or due to the storage conditions. Therefore, it is essential to characterize the structural heterogeneity of mAbs with respect to properties including conformational changes, aggregation and post-translational modifications. In this work, Zhu et al. at…
Important advances in top-down and middle-level analytical LC-MS strategies have arisen in recent years, focused on the characterization of therapeutic monoclonal antibodies. Similar strategies to analyze polyclonal IgG in regard to subclass abundance and glycosylation patterns may provide new insight into immune regulatory processes. However, challenges associated with molecular heterogeneity due to inherent sequence variability…
Scientists at the University of Amsterdam use SmartEnzymes in a new study of the biological and structural properties of antibody pathogenesis in blood transfusions. Blood transfusions are a vital part of healthcare but can in some cases lead to severe conditions such as anti-leukocyte antibodies in the transfusion product that cause transfusion-mediated acute lung…
Srzentic et al. present an interlaboratory study investigating the current state of top-down and middle-down mass spectrometry methods for the characterization of therapeutic monoclonal antibodies and explore the potential added value of these methods to the more commonly used bottom-up mass spectrometry. In the growing market of therapeutic monoclonal antibodies (mAbs), there is an…
Gstöttner et al. used SmartEnzymes for their CE-MS analysis of highly homologous bispecifics, one of them containing LALA mutations. Bispecific antibodies (BsAb) offer an attractive alternative to traditional mAbs since they can bind to two different epitopes simultaneously. Because they are composed of four different peptide chains, rather than two, they are subject to…
The Tn-antigen is characterized by the presence of a single N-Acetylgalactosamine (GalNAc) residue linked to serine or threonine residues on proteins. It is an immature glycoform without extension to form any of the common O-glycan core structures. The Tn antigen is a hallmark of many forms of cancer and rarely found in healthy tissue. Mapping and comparing Tn-antigens on proteins is therefore…
