Agilent Technologies, Inc.
Articles
Robust Oligonucleotide Molecular Weight Confirmation via LC/MS
October 31st 2024Oligonucleotides—therapeutic agents for difficult-to-treat diseases—rely on precise molecular weight confirmation to ensure quality during development. Experts from Agilent Technologies explore the challenges of liquid chromatography-mass spectrometry (LC/MS) analysis in oligonucleotide synthesis, as well as strategies for accurate deconvolution, method development, and optimal equipment selection.
Top-down characterization of engineered Bcl-xL proteoforms
October 11th 2024Top-down fragmentation enables rapid characterization of phosphorylated proteins without extensive sample preparation and digestion. In this study, electron capture dissociation (ECD) was used to fragment proteoforms of the cell death-related protein, Bcl-xL. Using these methods, 85–90% sequence coverage was achieved for Bcl-xL proteoforms, allowing for effective localization of phosphorylation within minutes.
Antibody peptide mapping using the new Agilent ExD cell
October 11th 2024Enhanced antibody analysis using electron capture dissociation (ECD) allows for precise glycan localization in low-abundance glycopeptides. This study compares the fragmentation of trastuzumab tryptic digests using ECD and collision-induced dissociation (CID). While CID generates abundant glycan HexNAc ions at 204 m/z, ECD preserves the labile glycan group, enabling accurate site localization.
Identification of Amino Acid Isomers Using Electron Capture Dissociation
October 11th 2024Electron capture dissociation produces distinct fragments of amino acid side chains, enabling the identification of isomeric amino acids such as leucine and isoleucine. This application note demonstrates the isomer identification workflow for peptides and intact proteins using the new Agilent ExD cell and ExDViewer software for fragment analysis.
Trends, Best Practices, and Analytical Challenges in Chemical Characterization of Medical Devices
October 7th 2024Chemical characterization of medical devices, including drug-device combination products, is crucial for ensuring regulatory compliance and patient safety by identifying and quantifying chemicals that may interact with the human body. This paper explores current trends, best practices, and regulatory developments in extractables and leachables (E&L) testing for medical devices.
Reliable and Objective Antibody Analysis with the New Agilent ProteoAnalyzer System
September 18th 2024This technical overview compares the characterization of NISTmAb using the ProteoAnalyzer with published NIST data obtained via traditional single capillary CE-SDS technology. The results show strong correlation with the reference data, confirming that the ProteoAnalyzer is a reliable tool for analytical workflows.
More Sensitive Quantification of PFAS by LC/MS with the Agilent 1260 Infinity II Hybrid Multisampler
August 6th 2024This application note demonstrates the use of the Agilent 1260 Infinity II Hybrid Multisampler for the analysis of per- and polyfluorinated alkyl substances (PFAS). The Hybrid Multisampler in feed injection mode was used with optimized Feed Speed for the sample injection to trap and enrich the compounds on the column. This avoided peak broadening and breakthrough of the early-eluting polar PFAS compounds. The less polar PFAS compounds can be enriched for more sensitive quantification.
Ultra-Trace Quantification of Per- and Polyfluoroalkyl Substances (PFAS) in Drinking Water
August 6th 2024The US EPA has released interim health advisory limits (HALs) of 4 and 20 pg/L in drinking water for the PFAS perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS), respectively. This application note addresses PFAS testing at these challenging new levels in aqueous samples using optimized solid-phase sample extraction and LC/MS/MS analysis with an Agilent 1290 Infinity II liquid chromatography (LC) system coupled to an Agilent 6495C triple quadrupole (TQ) mass spectrometer.
Protein Therapeutics: Antibody Characterization with ECD
June 10th 2024Labile post-translational modifications are better preserved using ECD than conventional MS fragmentation techniques such as collision-induced dissociation (CID). Common modifications, such as glycosylation and phosphorylation, can impact the safety, efficacy, and binding activity of monoclonal antibodies but are challenging to identify. Stephen Sciuto from Agilent Technologies and colleagues analyzed tryptic digests of NIST mAb and Infliximab using an 6545XT Q-TOF mass spectrometer in ExD or CID mode. ECD provided higher sequence coverage than CID, reaching 100% for glycosylated peptides in Infliximab. The ability to pinpoint glycosylation sites using fragment ions demonstrates the power of ECD for characterizing glycopeptides.
Monoclonal Antibodies: Faster Analysis with Microdroplets
June 10th 2024The first reported combination of microdroplet reactions with ECD fragmentation offers a quicker way to analyze monoclonal antibodies for disease treatments. Thomas Walker from Agilent Technologies and co-workers demonstrate the use of an Agilent Jet Stream electrospray ionization source to facilitate in-spray chemical reduction and protease digestion of monoclonal antibodies. Downstream fragmentation of the microdroplet reaction products with ECD provided rapid characterization of intact antibodies in minutes. Efficient ECD fragmentation yielded rich sequence information including evidence of disulfide bond connectivity and confirmation of key sequences called complementary determining regions. These results highlight this method’s potential for fast and cost-effective antibody characterization with minimal sample preparation.
Synthetic Peptides: Chromatographic Methods for Separation, Quantification, and Characterization
May 9th 2024Peptides are versatile molecules that have a wide range of applications in biotechnology, medicine, and research. They can be used as drugs, diagnostics, biomarkers, or tools to study cellular processes. However, peptides also pose unique challenges in terms of analysis and purification due to their complexity, diversity, and sensitivity.
