A5052968814- Glycosylation and Glycoproteins Research
- Monoclonal and Polyclonal Antibodies Research
- Protein purification and stability
- Galectins and Cancer Biology
- Analytical Chemistry and Chromatography
- Melanoma and MAPK Pathways
- Computational Drug Discovery Methods
- Carbohydrate Chemistry and Synthesis
- T-cell and B-cell Immunology
- Mass Spectrometry Techniques and Applications
- Metabolomics and Mass Spectrometry Studies
- Blood groups and transfusion
- HIV Research and Treatment
- Protein Kinase Regulation and GTPase Signaling
- Pharmacogenetics and Drug Metabolism
- Chemokine receptors and signaling
- Electrochemical Analysis and Applications
- Innovative Microfluidic and Catalytic Techniques Innovation
- Immunodeficiency and Autoimmune Disorders
- Immune Cell Function and Interaction
- Advanced Proteomics Techniques and Applications
- Receptor Mechanisms and Signaling
- Systemic Lupus Erythematosus Research
- Complement system in diseases
- Infant Nutrition and Health
Leiden University Medical Center
2016-2025
Netherlands Metabolomics Centre
2015-2025
Proteogenomics Research Institute for Systems Medicine
2015-2017
Vrije Universiteit Amsterdam
2010-2015
SC Solutions (United States)
2014
University of Münster
2009
Current approaches to study glycosylation of polyclonal human immunoglobulins G (IgG) usually imply protein digestion or glycan release. While these allow in-depth characterization, they also result in a loss valuable information regarding certain subclasses, allotypes and co-occuring post-translational modifications (PTMs). Unfortunately, the high variability IgGs makes their intact mass spectrometry (MS) analysis extremely challenging. We propose here middle-up strategy for fragment...
Monomeric serum immunoglobulin A (IgA) can contribute to the development of various autoimmune diseases, but regulation IgA effector functions is not well defined. Here, we show that two subclasses (IgA1 and IgA2) differ in their effect on immune cells due distinct binding signaling properties. Whereas IgA2 acts pro-inflammatory neutrophils macrophages, IgA1 does have pronounced effects. Moreover, different glycosylation profiles, with possessing more sialic acid than IgA2. Removal increases...
Antibody dependent cellular cytotoxicity (ADCC) is an Fc-dependent effector function of IgG important for anti-viral immunity and anti-tumor therapies. NK-cell mediated ADCC mainly triggered by IgG-subclasses IgG1 IgG3 through the IgG-Fc-receptor (FcγR) IIIa. Polymorphisms in immunoglobulin gamma heavy chain gene likely form a layer variation strength ADCC-response, but this has never been studied detail. We produced all 27 known allotypes assessed FcγRIIIa binding activity. While IgG1, IgG2...
To monitor the Fc glycosylation of therapeutic immunoglobulin G in bioprocess development, product characterization and release analytics, reliable techniques for analysis are needed. Several analytical methods suitable this application. We recently presented results comparing detection glycan that separation-based, but did not include mass spectrometry (MS). In study reported here, we comprehensively compared MS-based profiling an IgG biopharmaceutical. A antibody reference material was...
Bottom-up glycoproteomics by liquid chromatography–mass spectrometry (LC–MS) is an established approach for assessing glycosylation in a protein- and site-specific manner. Consequently, tools are needed to automatically align, calibrate, integrate LC–MS data. We developed modular software package designed tackle the individual aspects of experiment, called LaCyTools. Targeted alignment performed using user defined m/z retention time (tr) combinations. Subsequently, sum spectra created each...
Glycosylation is a common co- and post-translational protein modification, having large influence on properties like conformation solubility. Furthermore, glycosylation an important determinant of efficacy clearance biopharmaceuticals such as immunoglobulin G (IgG). Matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS) shows potential for the site-specific analysis IgG at glycopeptide level. With this approach, however, information about sialylation...
The in-depth, high-sensitivity characterization of the glycome from complex biological samples, such as biofluids and tissues, is utmost importance in basic research biomarker discovery. Major challenges often arise vast structural diversity glycans combination with limited sample amounts. Here, we present a method for highly sensitive released N-glycans by combining capillary electrophoresis-electrospray ionization-mass spectrometry (CE-ESI-MS) approach linkage-specific derivatization...
The study of N-linked glycosylation has long been complicated by a lack bioinformatics tools. In particular, there is still fast and robust data processing tools for targeted (relative) quantitation. We have developed modular, high-throughput software, MassyTools, that capable calibrating spectra, extracting data, performing quality control calculations based on user-defined list glycan or glycopeptide compositions. Typical examples output include relative areas after background subtraction,...
Glycosylation is a topic of intense current interest in the development biopharmaceuticals because it related to drug safety and efficacy. This work describes results an interlaboratory study on glycosylation Primary Sample (PS) NISTmAb, monoclonal antibody reference material. Seventy-six laboratories from industry, university, research, government, hospital sectors Europe, North America, Asia, Australia submitted total 103 reports glycan distributions. The principal objective this was...
Glycans expand the structural complexity of proteins by several orders magnitude, resulting in a tremendous analytical challenge when including them biomedical research. Recent glycobiological research is painting picture which glycans represent crucial and functional component majority proteins, with alternative glycosylation lipids being an important regulatory mechanism many biological pathological processes. Since interindividual differences are extensive, large studies needed to map...
Over the last years, numerous strategies have been proposed to enhance both ionization efficiency and spray stability in electrospray (ESI), particular for nanospray applications. In nano-liquid chromatography–mass spectrometry (nano-LC–ESI-MS), a better ESI performance has observed when coaxial gas flow is added around emitter. Moreover, enrichment of with an organic dopant led improved desolvation overall enhanced sensitivity. this study, use enriched nitrogen (DEN)-gas combined sheathless...
Abstract Pregnancy requires partial suppression of the immune system to ensure maternal-foetal tolerance. Protein glycosylation and especially terminal sialic acid linkages, are prime importance in regulating pro- anti-inflammatory responses. However, little is known about pregnancy-associated changes serum N -glycome linkages. Using a combination recently developed methods, i.e. derivatisation that allows distinction between α2,3- α2,6-linked acids by high-throughput MALDI-TOF-MS...
<h3>Objectives</h3> Patients with autoantibody-positive rheumatoid arthritis (RA) are less likely to experience pregnancy-induced improvement of RA disease activity (DAS28-C reactive protein (CRP)) compared patients autoantibody-negative RA. Anti-citrullinated antibodies (ACPAs) the most specific autoantibodies for We previously demonstrated that is associated changes in total IgG glycosylation, which regulate antibody effector function. Therefore, we sought analyse ACPA-IgG glycosylation...
Determination of the impact individual antibody glycoforms on FcɣRIIIa affinity, and consequently antibody-dependent cell-mediated cytotoxicity (ADCC) previously required high purity glycoengineering. We hyphenated affinity chromatography to mass spectrometry, which allowed direct comparison intact monoclonal antibodies. The approach enabled reproduction refinement known glycosylation effects, insights afucosylation pairing as well low-abundant, unstudied glycoforms. Our method greatly...
Immunoglobulin (Ig) glycosylation is recognized for its influence on Ig turnover and effector functions. However, the large-scale profiling of in a biomedical setting challenged by existence different isotypes subclasses, their varying serum concentrations, presence multiple sites per Ig. Here, high-throughput nanoliquid chromatography (LC)- mass spectrometry (MS)-based method simultaneous analysis IgG IgA glycopeptides was developed applied sample set from 185 healthy donors. Sample...
Broadly neutralizing antibodies (bNAbs) targeting the HIV-1 envelope glycoprotein (Env) have capacity to delay viral rebound when administered people with (PWH) during anti-retroviral therapy (ART) interruption. To further enhance performance of bNAbs through their Fc effector functions, in particular NK cell-mediated killing infected cells, we produced a panel glyco-engineered (afucosylated) enhanced affinity for gamma receptor IIIa. These afucosylated anti-HIV-1 cell activation and...
Good pharmacokinetic (PK) behavior is a key prerequisite for sufficient efficacy of therapeutic monoclonal antibodies (mAbs). Fc glycosylation critical quality attribute (CQA) mAbs, due to its impact on stability and effector functions. However, the effects various IgG glycoforms antibody PK remain unclear. We used combination glycoengineering glycoform-resolved measurements by mass spectrometry (MS) assess glycoform PK. Four differently glycoengineered each still containing multiple...
The asialoglycoprotein receptor (ASGPR) and the mannose C-type 1 (MRC1) are well known for their selective recognition clearance of circulating glycoproteins. Terminal galactose N-Acetylgalactosamine recognized by ASGPR, while terminal mannose, fucose, N-Acetylglucosamine MRC1. effects ASGPR MRC1 deficiency on N-glycosylation individual proteins have been studied. However, impact homeostasis major plasma glycoproteins is debated glycosylation has not mapped with high molecular resolution in...