Journal of the American Society for Mass Spectrometry最新文献

{"title":"Online Coupling of Asymmetrical Flow Field-Flow Fractionation with Native Mass Spectrometry for Monoclonal Antibody Characterization.","authors":"Yuetian Yan, Tao Xing, Shunhai Wang, Ning Li","doi":"10.1021/jasms.5c00089","DOIUrl":"https://doi.org/10.1021/jasms.5c00089","url":null,"abstract":"<p><p>Integration of Asymmetrical Flow Field-Flow Fractionation (AF4) with online native mass spectrometry (MS) is an attractive concept that offers significant value for the analytical characterization of protein therapeutics. Its execution, however, is challenged by the incompatibility of mobile phase components and the difficulties of achieving sensitive MS detection without compromising the separation efficiency. Herein, we describe in detail the successful coupling of a commercial AF4 instrument with online native MS detection. By optimizing the operational conditions of AF4 using ammonium acetate-based mobile phases and fine-tuning the interface configurations, the established AF4-MS platform achieved uncompromised separation performance while enabling sensitive online native MS measurements. The feasibility of the method for studying protein analytes was first demonstrated by analyzing a protein mixture sample (17 kDa to 660 kDa), followed by size variant characterization of the NISTmAb. The AF4-MS method was subsequently compared with native SEC-MS using both protein mixture and mAb samples, highlighting their complementarity as size-based separation techniques and showcasing the quantitative advantages of AF4-MS. Applications of AF4-MS in supporting therapeutic mAb development were demonstrated using two case studies, including heterodimer quantitation in a coformulated mAb cocktail sample and antibody-antigen binding stoichiometry characterization. This developed AF4-MS method is easily implementable and could be a valuable addition to our analytical toolbox to support therapeutic mAb development.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-06-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144264952","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Increasing Oligonucleotide Sequencing Information and Throughput with Ion Mobility Spectrometry-Mass Spectrometry.","authors":"Jack P Ryan, Gordon W Slysz, Peter Rye, Sarah M Stow, James N Dodds, John Sausen, Erin S Baker","doi":"10.1021/jasms.5c00083","DOIUrl":"https://doi.org/10.1021/jasms.5c00083","url":null,"abstract":"<p><p>Synthetic oligonucleotides, such as antisense oligonucleotides or small interfering RNA, are small chain nucleic acid polymers that can be used therapeutically to control gene expression. Watson-Crick base pair interactions provide the primary mode of interaction between synthetic oligonucleotides and their target molecule, and this binding requires accurate, robust, and rapid sequence verification. The development of high-quality synthetic oligonucleotides and comprehensive analytical workflows for their evaluation is therefore essential. Herein, a platform coupling liquid chromatography, ion mobility spectrometry, collision-induced dissociation, and mass spectrometry (LC-IMS-CID-MS) was applied to facilitate oligonucleotide sequence confirmation. Using IMS, multiple charge states of the same oligonucleotide were mobility separated and analyzed simultaneously. Furthermore, all-ion fragmentation was implemented to provide sequence coverage for each charge state using fewer injections than current targeted multiple injection LC-MS methods. The results demonstrated herein denote sequence coverage is generally inversely proportional to oligonucleotide length (<i>i</i>.<i>e</i>., lower fidelity coverage for longer strands), with observed sequence coverages ranging between 40% and 80% for molecules comprised of 20-40 residues. To ease the burden of spectral interpretation and sequence determination for the LC-IMS-CID-MS data, an analysis workflow using the Pacific Northwest National Laboratories (PNNL) preprocessor and Agilent's BioConfirm software was developed.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144257069","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Data-Independent Acquisition Parallel Accumulation-Serial Fragmentation (diaPASEF) Analysis of the Separated Zebrafish Lens Improves Identifications.","authors":"Sarah R Zelle, W Hayes McDonald, Kristie L Rose, Hassane S Mchaourab, Kevin L Schey","doi":"10.1021/jasms.5c00087","DOIUrl":"https://doi.org/10.1021/jasms.5c00087","url":null,"abstract":"<p><p>Ocular lens fiber cells degrade their organelles during differentiation to prevent light scattering. Organelle degradation occurs continuously throughout an individual's lifespan, creating a spatial gradient of young cortical fiber cells in the lens periphery to older nuclear fiber cells in the center of the lens. Therefore, separation of cortical and nuclear regions enables examination of protein aging. Previously, the human lens cortex and nucleus have been studied using data-independent acquisition (DIA) proteomics, allowing for the identification of low-abundance protein groups. In this study, we employed data-independent acquisition parallel accumulation-serial fragmentation (diaPASEF) proteomics on a timsTOF HT instrument to study the zebrafish lens proteome and compared results to a standard DIA method employed on an Orbitrap Exploris 480 mass spectrometer. Using the additional ion mobility gas phase separation of diaPASEF, peptide and protein group identifications increased by over 200% relative to an Orbitrap DIA method in the zebrafish lens. With diaPASEF, we identified 13,721 and 11,996 unique peptides in the cortex and nucleus of the zebrafish lens, respectively, which correspond to 1,537 and 1,389 protein groups. Thus, separation of the zebrafish lens into cortical and nuclear regions followed by diaPASEF analysis produced the most comprehensive zebrafish lens proteomic data set to date.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144257068","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development and Characterization of a Fluorinated MS-Cleavable Cross-Linker for Structural Proteomics","authors":"Oleksandr Sorokin,&nbsp;Frank Hause,&nbsp;Christian H. Ihling,&nbsp;Tomáš Vranka,&nbsp;Václav Matoušek and Andrea Sinz*,&nbsp;","doi":"10.1021/jasms.4c0048910.1021/jasms.4c00489","DOIUrl":"https://doi.org/10.1021/jasms.4c00489https://doi.org/10.1021/jasms.4c00489","url":null,"abstract":"<p >Cross-linking mass spectrometry (XL-MS) is an important method for studying three-dimensional protein structures and mapping protein–protein interactions. Some limitations of XL-MS still consist of its use for in-cell and in vivo applications. To date, cross-linking reagents are urgently needed that can efficiently penetrate the cell membrane to comprehensively map protein–protein interaction networks in intact cells. In this study, the fluorinated MS-cleavable cross-linker bis(pentafluorophenyl) ureido-4,4′-dibutyrate (DPFU) is described. DPFU is based on the MS-cleavable cross-linker disuccinimidyl dibutyric urea (DSBU) with the aim of balancing the hydrophobicity and solubility to improve membrane permeability. DPFU was evaluated for its solubility behavior in different detergent solutions to optimize conditions for its potential application in live cells. Using bovine serum albumin (BSA) as a model protein, XL-MS experiments were conducted across different temperatures and cross-linker concentrations. Solubility assays identified sodium dodecyl sulfate (SDS) as effective for enhancing DPFU solubility in an aqueous environment. DPFU yielded fewer cross-links for BSA than DSBU, highlighting limitations regarding its cross-linking efficiency under similar experimental conditions. This study provides the first insights into fluorinated cross-linkers, suggesting that further optimization is needed for a broader application of DPFU for future in-cell XL-MS studies.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":"36 6","pages":"1410–1413 1410–1413"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/epdf/10.1021/jasms.4c00489","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144202562","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Faces of Mass Spectrometry/Richard Y.-C. Huang.","authors":"Anne Brenner, J D Brookbank","doi":"10.1021/jasms.5c00097","DOIUrl":"10.1021/jasms.5c00097","url":null,"abstract":"","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":"1204-1207"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143957624","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"LC-ESI(±)-LTQ MSⁿ-Based Metabolomic Profiling of Coffee: Fragmentation Pathways for Identification of Major Polar Compounds.","authors":"Marcos Valério Vieira Lyrio, Danieli Grancieri Debona, Amanda Eiriz Feu, Nayara Araujo Dos Santos, Arlan da Silva Gonçalves, Ricardo Machado Kuster, Eustáquio Vinícius Ribeiro de Castro, Wanderson Romão","doi":"10.1021/jasms.4c00418","DOIUrl":"10.1021/jasms.4c00418","url":null,"abstract":"<p><p>Coffee is characterized by a complex chemical matrix that significantly influences its organoleptic properties and market value. This complexity is driven by factors such as botanical species, geographical origin, cultivation conditions, and post-harvest processing methods. Metabolomic studies aim to elucidate how these factors impact the biosynthesis of metabolites that contribute to the sensory qualities of high-quality coffee. Among various analytical techniques, liquid chromatography-mass spectrometry (LC-MS) is particularly effective for separating, identifying, and quantifying these compounds. Most metabolomic studies employ high-resolution mass spectrometry (HRMS) for its superior mass accuracy (<1 ppm), whereas the interpretation of low-resolution data requires additional effort, often relying on literature references and proposed fragmentation mechanisms. In this study, we applied LC-ESI(±)LTQ MSⁿ to comprehensively profile coffee metabolites, identifying 60 compounds, including polar compounds and their isomers such as chlorogenic acids, carbohydrates, amino acids, alkaloids, glycosylated diterpenes, and flavonoids. Fragmentation mechanisms were proposed and discussed. The results demonstrate the effectiveness of LC-ESI(±)LTQ MSⁿ in a detailed metabolomic analysis, providing a robust platform for future research in coffee metabolomics.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":"1213-1226"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12142669/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143957004","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development and Characterization of a Fluorinated MS-Cleavable Cross-Linker for Structural Proteomics.","authors":"Oleksandr Sorokin, Frank Hause, Christian H Ihling, Tomáš Vranka, Václav Matoušek, Andrea Sinz","doi":"10.1021/jasms.4c00489","DOIUrl":"10.1021/jasms.4c00489","url":null,"abstract":"<p><p>Cross-linking mass spectrometry (XL-MS) is an important method for studying three-dimensional protein structures and mapping protein-protein interactions. Some limitations of XL-MS still consist of its use for in-cell and in vivo applications. To date, cross-linking reagents are urgently needed that can efficiently penetrate the cell membrane to comprehensively map protein-protein interaction networks in intact cells. In this study, the fluorinated MS-cleavable cross-linker bis(pentafluorophenyl) ureido-4,4'-dibutyrate (DPFU) is described. DPFU is based on the MS-cleavable cross-linker disuccinimidyl dibutyric urea (DSBU) with the aim of balancing the hydrophobicity and solubility to improve membrane permeability. DPFU was evaluated for its solubility behavior in different detergent solutions to optimize conditions for its potential application in live cells. Using bovine serum albumin (BSA) as a model protein, XL-MS experiments were conducted across different temperatures and cross-linker concentrations. Solubility assays identified sodium dodecyl sulfate (SDS) as effective for enhancing DPFU solubility in an aqueous environment. DPFU yielded fewer cross-links for BSA than DSBU, highlighting limitations regarding its cross-linking efficiency under similar experimental conditions. This study provides the first insights into fluorinated cross-linkers, suggesting that further optimization is needed for a broader application of DPFU for future in-cell XL-MS studies.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":"36 6","pages":"1410-1413"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12142660/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144214586","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Spatial Metabolomics Annotation Workflow Leveraging Cyclic Ion Mobility and Machine Learning-Predicted Collision Cross Sections.","authors":"Dmitry Leontyev, Eric C Gier, Viraj A Master, Rebecca S Arnold, John A Petros, Facundo M Fernández","doi":"10.1021/jasms.5c00090","DOIUrl":"10.1021/jasms.5c00090","url":null,"abstract":"<p><p>In nontargeted spatial metabolomics, accurate annotation is crucial for understanding metabolites' biological roles and spatial patterns. MS² mass spectrometry imaging (MSI) coverage is often incomplete or nonexistent, resulting in many unknown features that represent an untapped source of biological information. Ion mobility-derived collision cross sections (CCS) have been leveraged as valuable descriptors for confirming putative metabolite annotations, distinguishing isomers, and aiding in unknown structural elucidation. In this study, desorption electrospray ionization cyclic ion mobility mass spectrometry imaging (DESI-cIM-MSI) data from human renal cell carcinoma (RCC) tissues is used as a testbed to explore the extent to which CCS measurements enhance MSI lipid annotation confidence when combined with machine learning CCS predictions and SIRIUS analysis of MS² data. Multipass IM experiments yielded excellent CCS accuracy (<0.4%) relative to database values for differential lipids found in RCC tissues, improving the filtering threshold used in previous CCS-based annotation workflows. High-accuracy multipass CCS measurements enabled the correct annotation of isobaric lipid database matches, even in the absence of MS² data. Additionally, MS² data from differential RCC features were uploaded to SIRIUS, and the predicted CCS values for SIRIUS candidates were compared to experimental CCS data to filter out unlikely candidates. Finally, CCS measurements contributed to the annotation of two spatially correlated unknown features, differential between tumor and control kidney tissues. Both features were assigned to rocuronium, a surgical muscle relaxant that had not been previously reported in MSI studies. Overall, these results underscore the potential of high-accuracy CCS values to enhance metabolite annotations in MSI-based spatial metabolomics.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":"1386-1394"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12142677/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144109379","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Dual Apodization Maximizes Charge Resolution and Frequency Precision in Charge Detection Mass Spectrometry.","authors":"Raj A Parikh, Martin F Jarrold","doi":"10.1021/jasms.5c00049","DOIUrl":"10.1021/jasms.5c00049","url":null,"abstract":"<p><p>Charge detection mass spectrometry (CD-MS) is a single-particle technique in which the masses of individual ions are determined from simultaneous measurements of their <i>m</i>/<i>z</i> ratio and charge. Ions are trapped in an electrostatic linear ion trap and oscillate back and forth through a detection cylinder coupled to a low noise charge sensitive amplifier. The resulting signal is analyzed using short-time Fourier transforms (STFTs) to determine the <i>m</i>/<i>z</i> ratio and charge. The <i>m</i>/<i>z</i> ratio is determined from the oscillation frequency, and the charge is obtained from the magnitude of the fundamental. Here we compare the methods used to analyze time domain data for single ion measurements including STORI plots. We conclude that the original STFT approach remains the best method for the analysis of CD-MS data. However, there are many ways of implementing the STFT approach. We compare the options with the goal of maximizing precision of the charge and <i>m</i>/<i>z</i> determinations while simultaneously maximizing the number of ions that are detected. A variety of apodization methods are compared, and the effects of scalloping loss, equivalent noise bandwidth, computation time, window length, and step size are evaluated. Maximizing the precision of the charge and <i>m</i>/<i>z</i> determinations places conflicting constraints on the window length, and we conclude that a dual apodization strategy, with different window lengths, provides the most robust approach to analyzing results for the broad range of different samples that can be measured by CD-MS.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":"1325-1335"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143956761","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Rapid Assessment of Samples from Large-Scale Clandestine Synthetic Drug Laboratories by Soft Ionization by Chemical Reaction in Transfer-High-Resolution Mass Spectrometry.","authors":"Maximilian Greif, Tobias Frömel, Thomas P Knepper, Carolin Huhn, Stephan Wagner, Michael Pütz","doi":"10.1021/jasms.5c00006","DOIUrl":"10.1021/jasms.5c00006","url":null,"abstract":"<p><p>The worldwide ongoing trend of synthetic drug production is also of increasing concern due to enormous amounts of chemical waste produced in clandestine laboratories. Typically, several tons of different types of production waste are stored in numerous containers and need to be characterized after dismantling a laboratory to assess production features, e.g., synthesis route and production scale, and to draw conclusions on the minimum number of batches produced. This forensic assessment is commonly done by a rather laborious gas chromatography - mass spectrometry approach. The aim of this work is to evaluate the suitability of the SICRIT (soft ionization by chemical reaction in transfer) ion source, which is based on the dielectric barrier discharge ionization principle, combined with high-resolution mass spectrometry (HRMS), for the rapid classification of liquid samples from amphetamine production in a seized large-scale clandestine drug laboratory. Among the different sample introduction methods tested, headspace analysis directly into the SICRIT ion source in conjunction with a heated inlet proved to be optimal. Identification of expected target substances (reaction educts, intermediates, byproducts, products) was possible as well as grouping related samples and assigning them to specific synthesis steps by multivariate data analysis in an unsupervised approach. In addition, supervised machine learning algorithms were evaluated to obtain a classification model for the assessment of production waste samples from one dismantled synthetic drug laboratory, and a random forest classifier showed the best performance with an accuracy of 97%. The potential of the novel SICRIT-HRMS approach for the assessment of synthetic drug laboratories was demonstrated.</p>","PeriodicalId":672,"journal":{"name":"Journal of the American Society for Mass Spectrometry","volume":" ","pages":"1254-1263"},"PeriodicalIF":3.1,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143958757","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}