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Enhanced Discovery of Alternative Proteins (AltProts) in Mouse Cardiac Development Using Data-Independent Acquisition (DIA) Proteomics.
Anal Chem
January 2025
Department of Applied Biology and Chemical Technology, State Key Laboratory of Chemical Biology and Drug Discovery, Hong Kong Polytechnic University, Hong Kong 999077, China.
Alternative proteins (AltProts) are a class of proteins encoded by DNA sequences previously classified as noncoding. Despite their historically being overlooked, recent studies have highlighted their widespread presence and distinctive biological roles. So far, direct detection of AltProt has been relying on data-dependent acquisition (DDA) mass spectrometry (MS).
View Article and Find Full Text PDFUHPLC-TIMS-PASEF-MS for Lipidomics: From Theory to Practice.
Methods Mol Biol
January 2025
Department of Medicine and Surgery, Proteomics and Metabolomics Unit, University of Milano-Bicocca, Vedano al Lambro, Italy.
Trapped ion mobility spectrometry (TIMS) using parallel accumulation serial fragmentation (PASEF) is an advanced analytical technique that offers several advantages in mass spectrometry (MS)-based lipidomics. TIMS provides an additional dimension of separation to mass spectrometry and accurate collision cross-section (CCS) measurements for ions, aiding in the structural characterization of molecules. This is especially valuable in lipidomics for identifying and distinguishing isomeric or structurally similar compounds.
View Article and Find Full Text PDFA Protocol for GC-MS Profiling of Chiral Secondary Amino Acids.
Methods Mol Biol
January 2025
Laboratory of Analytical Biochemistry & Metabolomics, Biology Centre, Czech Academy of Sciences, České Budějovice, Czech Republic.
A simple analytical workflow is described for gas chromatographic-mass spectrometric (GC-MS)-based chiral profiling of secondary amino acids (AAs) in biological matrices. The sample preparation is carried out directly in aqueous biological sample extracts and involves in situ heptafluorobutyl chloroformate (HFBCF) derivatization-liquid-liquid microextraction of nonpolar products into hexane phase followed by subsequent formation of the corresponding methylamides from the HFB esters by direct treatment with methylamine reagent solution. The (O, N) HFB-butoxycarbonyl-methylamide AA products (HFBOC-MA) are separated on a Chirasil-L-Val capillary column and quantitatively measured by GC-MS operated in selected ion monitoring (SIM) mode.
View Article and Find Full Text PDFQuality Control and Validation Issues in LC-MS-Based Metabolomics.
Methods Mol Biol
January 2025
Division of Systems Medicine, Department of Metabolism Department of Metabolism, Digestion and Reproduction, Imperial College, London, UK.
Metabolic profiling performed using untargeted metabolomics of different, complex biological samples aims to apply agnostic/holistic, hypothesis-free, analysis of the small molecules that are present in the analyzed sample. This approach has been the center of major investments and dedicated efforts from the research community for many years. However, limitations and challenges remain, particularly with regard to the validation and the quality of the obtained results.
View Article and Find Full Text PDFQuality Assurance in Metabolomics and Metabolic Profiling.
Methods Mol Biol
January 2025
Center for Environmental Measurement and Modeling, Environmental Protection Agency, Athens, GA, USA.
Metabolic profiling (untargeted metabolomics) aims for a global unbiased analysis of metabolites in a cell or biological system. It remains a highly useful research tool used across various analytical platforms. Incremental improvements across multiple steps in the analytical process may have large consequences for the end quality of the data.
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