Assessment of product quality attributes such as charge heterogeneity is an upmost requisite for the release of a monoclonal antibody (mAb). Analytical techniques, such as cation-exchange chromatography (CEX), accomplish this, causing the mAb to separate into acidic, main species, and basic variants. Here, an online volatile-salt-containing two-dimensional liquid chromatography (2D-LC) method coupled with mass spectrometry (MS) was performed to characterize the charge heterogeneity of mAbs using CEX chromatography in the first dimension (D) and anion-exchange chromatography (AEX) in the second dimension (D). The main peak of the CEX profile of D was transferred through a 2D heart-cut method to D for further analysis by the AEX-MS method. In the CEX method, mAb A showed 10 distinct variants, while the AEX method resulted in eight variants. However, a total of 13 variants were successfully resolved for mAb A in the 2D method. Similarly, mAb B exhibited seven variants in the CEX method and four variants in the AEX method, but the 2D-LC method revealed a total of nine variants for mAb B. Likewise, mAb C displayed seven variants in CEX and seven variants in AEX, whereas the 2D-LC method unveiled a total of 11 variants for mAb C. Additionally, native MS analysis revealed that the resolved charge variants were identified as amidation, oxidation, and isomerization of Asp variants in the main peak, which were not resolved in stand-alone methods. The present study demonstrates how 2D-LC can assist in identifying minor variations in charge distribution or conformation of mAb variants that would otherwise not be picked up by a single analytical method alone.
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http://dx.doi.org/10.1021/jasms.3c00308 | DOI Listing |
Clin Epigenetics
January 2025
Faculty of Medicine of TUD Dresden University of Technology, Institute for Clinical Genetics, University Hospital Carl Gustav Carus at TUD Dresden University of Technology, Dresden, Germany.
Autosomal dominant CDK13-related disease is characterized by congenital heart defects, dysmorphic facial features, and intellectual developmental disorder (CHDFIDD). Heterozygous pathogenic variants, particularly missense variants in the kinase domain, have previously been described as disease causing. Using the determination of a methylation pattern and comparison with an established episignature, we reveal the first hypomorphic variant in the kinase domain of CDK13, leading to a never before described autosomal recessive form of CHDFIDD in a boy with characteristic features.
View Article and Find Full Text PDFSignal Transduct Target Ther
January 2025
NHC Key Laboratory of Systems Biology of Pathogens, State Key Laboratory of Respiratory Health and Multimorbidity, National Institute of Pathogen Biology, and Center for Tuberculosis Research, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100730, China.
The global spread of Severe Acute Respiratory Syndrome Coronavirus 2. (SARS-CoV-2) and its variant strains, including Alpha, Beta, Gamma, Delta, and now Omicron, pose a significant challenge. With the constant evolution of the virus, Omicron and its subtypes BA.
View Article and Find Full Text PDFObjective: To test whether messenger RNA (mRNA) splicing is altered in neutrophils from patients with systemic lupus erythematosus (SLE) and can produce neoantigens.
Methods: RNA sequencing of neutrophils from patients with SLE (n = 15) and healthy donors (n = 12) were analyzed for mRNA splicing using the RiboSplitter pipeline, an event-focused tool based on SplAdder with subsequent translation and protein domain annotation. RNA sequencing from SARS-CoV2-infected individuals was used as an additional comparator.
Clin Genet
January 2025
Laboratorio de Citometría y Biología Molecular, Hospital de Clínicas, Universidad de la República, Montevideo, Uruguay.
Timeline and genetic analysis of a 55-year-old female with a family history of gastric cancer and multiple myeloma, who was diagnosed with AML and a germline CEBPA variant.
View Article and Find Full Text PDFJ Cyst Fibros
January 2025
Department NEUROFARBA, University of Florence; Paediatric and Liver Unit, Meyer Children's Hospital IRCCS, Florence, Italy.
Background: Elexacaftor-tezacaftor-ivacaftor (ETI) has significantly improved the clinical course of people with cystic fibrosis (pwCF) and eligible CFTR variants. In this study, we prospectively evaluated liver elastography, liver fibrosis indices and liver tests in children with CF aged 6-12 years started on ETI therapy.
Methods: Body mass index, sweat test, percent predicted forced expiratory volume in one second, serum markers of liver injury or portal hypertension, liver fibrosis indices, controlled attenuation parameter and liver stiffness were assessed before starting ETI and three and twelve months post-ETI, according to new international guidelines.
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