{"title":"From sensing to intelligence: A review of microfluidic-integrated optical and impedance platforms for single-cell analysis","authors":"Liangzun Fu,Jin Chen,Yang Zhang,Zongyi Wang,Xiwei Huang","doi":"10.1016/j.aca.2026.346347","DOIUrl":"https://doi.org/10.1016/j.aca.2026.346347","url":null,"abstract":"","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"64 1","pages":"346347"},"PeriodicalIF":6.2,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149091829","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}
Pub Date : 2026-09-29DOI: 10.1016/j.aca.2026.346333
José Raúl Belmonte-Sánchez, Roberto Romero-González, Antonia Garrido Frenich
Artificial intelligence (AI) has moved from a research topic to an operating tool across the chemical sciences, where large language models (LLMs), generative deep learning and agentic systems are now embedded in drug discovery, materials design and mass spectrometry (MS)-based analysis at a pace that has outstripped the peer-reviewed literature. Based on studies published between 2021 and 2026, this critical review argues that AI extends a recent lineage of disruptive tools in chemistry. It further proposes the human-in-the-loop agentic system, in which the chemist orchestrates and validates while the agent executes, as a candidate operative architecture for the coming decade, in preference to autonomous AI. Three converging streams (LLMs as scientific instruments, generative models for chemical design, and agentic systems with tool use and planning) are mapped onto a four-level spectrum of autonomy, from the single-LLM copilot to the autonomous robotic chemist. Practical implementation is illustrated through compound annotation in non-targeted MS, LLM-driven spectroscopic copilots, retrieval-augmented knowledge graphs, and AI coding assistants in routine methods development. On the debates over AI-assisted writing, publisher policies and peer review, it has been argued that transparent disclosure is a more robust path than detection or prohibition. It concludes by examining a possible productivity bifurcation between AI-adopting and AI-resistant groups by 2030 and proposing AI literacy as a core competency for analytical chemists in training.
{"title":"Human-in-the-loop AI in analytical chemistry: a critical review from large language models to agentic systems","authors":"José Raúl Belmonte-Sánchez, Roberto Romero-González, Antonia Garrido Frenich","doi":"10.1016/j.aca.2026.346333","DOIUrl":"https://doi.org/10.1016/j.aca.2026.346333","url":null,"abstract":"Artificial intelligence (AI) has moved from a research topic to an operating tool across the chemical sciences, where large language models (LLMs), generative deep learning and agentic systems are now embedded in drug discovery, materials design and mass spectrometry (MS)-based analysis at a pace that has outstripped the peer-reviewed literature. Based on studies published between 2021 and 2026, this critical review argues that AI extends a recent lineage of disruptive tools in chemistry. It further proposes the human-in-the-loop agentic system, in which the chemist orchestrates and validates while the agent executes, as a candidate operative architecture for the coming decade, in preference to autonomous AI. Three converging streams (LLMs as scientific instruments, generative models for chemical design, and agentic systems with tool use and planning) are mapped onto a four-level spectrum of autonomy, from the single-LLM copilot to the autonomous robotic chemist. Practical implementation is illustrated through compound annotation in non-targeted MS, LLM-driven spectroscopic copilots, retrieval-augmented knowledge graphs, and AI coding assistants in routine methods development. On the debates over AI-assisted writing, publisher policies and peer review, it has been argued that transparent disclosure is a more robust path than detection or prohibition. It concludes by examining a possible productivity bifurcation between AI-adopting and AI-resistant groups by 2030 and proposing AI literacy as a core competency for analytical chemists in training.","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"12 1","pages":""},"PeriodicalIF":6.2,"publicationDate":"2026-09-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149087382","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}
Pub Date : 2026-09-28DOI: 10.1016/j.aca.2026.346325
Ketolly N.S. Leal, Alan J. Stewart, Carlos H.I. Ramos, Marco A.Z. Arruda
Single-cell inductively coupled plasma mass spectrometry (SC-ICP-MS) has become an important tool for measuring elemental content in individual cells and revealing heterogeneity that is lost in bulk analysis. However, the reliability of these measurements is not determined by instrumental sensitivity alone. In practice, each stage of the workflow, from sample preparation and aerosol generation to transport efficiency, transient signal acquisition, and event discrimination, can alter the number of detected events and the calculated elemental mass per cell. This review discusses SC-ICP-MS from this workflow-centered perspective, focusing on the analytical bias introduced during sample handling, cell transport, instrumental acquisition, and data treatment. Attention is given to microflow introduction systems, collision/reaction cell strategies, ICP-MS/MS, ICP-TOF-MS, dwell-time selection, and Gaussian- and Poisson-based approaches for threshold definition. We also discuss how these factors affect the interpretation of endogenous elements, exogenous tracers, metal-labelled biomolecules, and particulate species in biological systems. Rather than treating single-cell distributions as direct representations of intracellular composition, we argue that the analytical workflow used to generate them should be considered when interpreting the data. This perspective is essential for improving comparability between studies and for moving SC-ICP-MS towards more reliable functional single-cell metallomics.
{"title":"Single-Cell ICP-MS: A Review of Analytical Bias Across the Workflow","authors":"Ketolly N.S. Leal, Alan J. Stewart, Carlos H.I. Ramos, Marco A.Z. Arruda","doi":"10.1016/j.aca.2026.346325","DOIUrl":"https://doi.org/10.1016/j.aca.2026.346325","url":null,"abstract":"Single-cell inductively coupled plasma mass spectrometry (SC-ICP-MS) has become an important tool for measuring elemental content in individual cells and revealing heterogeneity that is lost in bulk analysis. However, the reliability of these measurements is not determined by instrumental sensitivity alone. In practice, each stage of the workflow, from sample preparation and aerosol generation to transport efficiency, transient signal acquisition, and event discrimination, can alter the number of detected events and the calculated elemental mass per cell. This review discusses SC-ICP-MS from this workflow-centered perspective, focusing on the analytical bias introduced during sample handling, cell transport, instrumental acquisition, and data treatment. Attention is given to microflow introduction systems, collision/reaction cell strategies, ICP-MS/MS, ICP-TOF-MS, dwell-time selection, and Gaussian- and Poisson-based approaches for threshold definition. We also discuss how these factors affect the interpretation of endogenous elements, exogenous tracers, metal-labelled biomolecules, and particulate species in biological systems. Rather than treating single-cell distributions as direct representations of intracellular composition, we argue that the analytical workflow used to generate them should be considered when interpreting the data. This perspective is essential for improving comparability between studies and for moving SC-ICP-MS towards more reliable functional single-cell metallomics.","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"150 1","pages":""},"PeriodicalIF":6.2,"publicationDate":"2026-09-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149087556","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}
Pub Date : 2026-09-02DOI: 10.1016/j.aca.2026.346211
Katia Buonasera, Maurilio Galletta, Gianni Pezzotti Escobar, Simone Failla, Maria José Lo Faro, Francesco Priolo, Giovanna De Luca, Barbara Fazio, Antonio Alessio Leonardi, Alessia Irrera
Electrochemical surface-enhanced Raman spectroscopy (EC-SERS) is an advanced analytical technique that integrates the molecular specificity and high sensitivity of surface-enhanced Raman scattering with the dynamic control of electrochemistry. This hybrid approach enables real-time investigation of interfacial processes, including adsorption phenomena, charge-transfer interactions, and redox reactions at electrified surfaces. In recent years, significant progress has been achieved through the development of engineered plasmonic substrates, hybrid nanomaterials, and improved spectroelectrochemical configurations, greatly enhancing sensitivity, stability, and analytical performance.
{"title":"Electrochemical control meets enhanced Raman sensitivity: a critical review of electrochemical surface-enhanced Raman spectroscopy (EC-SERS) sensors","authors":"Katia Buonasera, Maurilio Galletta, Gianni Pezzotti Escobar, Simone Failla, Maria José Lo Faro, Francesco Priolo, Giovanna De Luca, Barbara Fazio, Antonio Alessio Leonardi, Alessia Irrera","doi":"10.1016/j.aca.2026.346211","DOIUrl":"https://doi.org/10.1016/j.aca.2026.346211","url":null,"abstract":"Electrochemical surface-enhanced Raman spectroscopy (EC-SERS) is an advanced analytical technique that integrates the molecular specificity and high sensitivity of surface-enhanced Raman scattering with the dynamic control of electrochemistry. This hybrid approach enables real-time investigation of interfacial processes, including adsorption phenomena, charge-transfer interactions, and redox reactions at electrified surfaces. In recent years, significant progress has been achieved through the development of engineered plasmonic substrates, hybrid nanomaterials, and improved spectroelectrochemical configurations, greatly enhancing sensitivity, stability, and analytical performance.","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"202 1","pages":""},"PeriodicalIF":6.2,"publicationDate":"2026-09-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148883960","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}
Pub Date : 2026-09-01DOI: 10.1016/j.aca.2026.346204
Cushla McGoverin
Vibrational spectroscopic methods, mid- and near-infrared, and Raman spectroscopies, have been extensively applied to the study of microorganisms. This is due to the relatively rapid analysis time, relative low-cost, experimental flexibility, and phenotypic nature of these methods. Consequently, vibrational spectroscopies have been investigated for clinical, environmental and industrial microbiology applications. This review outlines the principles of mid- and near-infrared, and Raman spectroscopies, and steps through vibrational spectroscopic experiments in the context of microbiological studies. Further applications within species/strain identification, antimicrobial resistance, bioremediation, cell sorting and intracellular microbial protein analysis are examined. Vibrational spectroscopy is expected to become increasingly routine in microbial analyses, particularly in clinical settings, driven by the harmonisation of spectral databases, the expanding application of deep learning methods and improved instrumentation to reduce sampling-to-reporting time.
{"title":"Vibrational spectroscopy of microorganisms: A review","authors":"Cushla McGoverin","doi":"10.1016/j.aca.2026.346204","DOIUrl":"https://doi.org/10.1016/j.aca.2026.346204","url":null,"abstract":"Vibrational spectroscopic methods, mid- and near-infrared, and Raman spectroscopies, have been extensively applied to the study of microorganisms. This is due to the relatively rapid analysis time, relative low-cost, experimental flexibility, and phenotypic nature of these methods. Consequently, vibrational spectroscopies have been investigated for clinical, environmental and industrial microbiology applications. This review outlines the principles of mid- and near-infrared, and Raman spectroscopies, and steps through vibrational spectroscopic experiments in the context of microbiological studies. Further applications within species/strain identification, antimicrobial resistance, bioremediation, cell sorting and intracellular microbial protein analysis are examined. Vibrational spectroscopy is expected to become increasingly routine in microbial analyses, particularly in clinical settings, driven by the harmonisation of spectral databases, the expanding application of deep learning methods and improved instrumentation to reduce sampling-to-reporting time.","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"14 1","pages":""},"PeriodicalIF":6.2,"publicationDate":"2026-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148883897","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}
Pub Date : 2026-09-01DOI: 10.1016/j.aca.2026.346212
Jaime Martínez-García,Clara de Lorenzo,Beatriz Fernández,Rosario Pereiro
{"title":"Biomedical and Clinical Frontiers of LA-ICP-MS: Insights from Cell Cultures and Animal Models to Human Samples","authors":"Jaime Martínez-García,Clara de Lorenzo,Beatriz Fernández,Rosario Pereiro","doi":"10.1016/j.aca.2026.346212","DOIUrl":"https://doi.org/10.1016/j.aca.2026.346212","url":null,"abstract":"","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"192 1","pages":"346212"},"PeriodicalIF":6.2,"publicationDate":"2026-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148895292","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}
Accurate detection and discrimination of glutathione (GSH) are critical for food safety and quality control. However, developing highly selective assays for the precise detection of GSH and its discrimination from other reducing substances remains a significant challenge. Herein, a promotion-effect-driven, dual-readout sensor array based on spindle-shaped Prussian blue (s-PB) nanozymes was presented for the highly selective detection and intelligent discrimination of GSH among various food additives. s-PB were synthesized via a simple double decomposition reaction followed by centrifugation and vacuum drying. The Fe3+/Fe2+ redox centers within the s-PB framework served as abundant peroxidase-like active sites, which catalyze the H2O2-mediated oxidation of ABTS to produce a characteristic green product. Interestingly, the addition of GSH (0 ∼ 625 μM) induced a concentration-dependent enhancement of both the absorbance intensity and color intensity, yielding a wide linear range (0.0125 ∼ 62.5 μM) and an ultralow detection limit of 0.013 μM. Leveraging s-PB as a single sensing receptor, a three-channel portable array was developed to accurately detect GSH and identify other food additives. The integration of smartphone imaging and machine learning algorithms enabled effective differentiation of food additives at various concentrations, mixtures using the proposed array. This work not only provided a highly sensitive and selective method for the "turn-on" detection of GSH, but also demonstrated a robust strategy for the multi-component analysis of food additives, thereby expanding the horizons of nanozymes in food safety monitoring.
{"title":"Overcoming the ''turn-off'' limitation via steric hindrance: A ''turn-on'' dual-readout nanozyme array based on spindle-shaped Prussian blue for highly selective discrimination of glutathione and food additives","authors":"Mei Li, Tianhui Huang, SuXuan Liao, Hailin Zhang, Jiakuan Wu, Qiuping Huang","doi":"10.1016/j.aca.2026.345668","DOIUrl":"10.1016/j.aca.2026.345668","url":null,"abstract":"<div><div>Accurate detection and discrimination of glutathione (GSH) are critical for food safety and quality control. However, developing highly selective assays for the precise detection of GSH and its discrimination from other reducing substances remains a significant challenge. Herein, a promotion-effect-driven, dual-readout sensor array based on spindle-shaped Prussian blue (s-PB) nanozymes was presented for the highly selective detection and intelligent discrimination of GSH among various food additives. s-PB were synthesized via a simple double decomposition reaction followed by centrifugation and vacuum drying. The Fe<sup>3+</sup>/Fe<sup>2+</sup> redox centers within the s-PB framework served as abundant peroxidase-like active sites, which catalyze the H<sub>2</sub>O<sub>2</sub>-mediated oxidation of ABTS to produce a characteristic green product. Interestingly, the addition of GSH (0 ∼ 625 μM) induced a concentration-dependent enhancement of both the absorbance intensity and color intensity, yielding a wide linear range (0.0125 ∼ 62.5 μM) and an ultralow detection limit of 0.013 μM. Leveraging s-PB as a single sensing receptor, a three-channel portable array was developed to accurately detect GSH and identify other food additives. The integration of smartphone imaging and machine learning algorithms enabled effective differentiation of food additives at various concentrations, mixtures using the proposed array. This work not only provided a highly sensitive and selective method for the \"turn-on\" detection of GSH, but also demonstrated a robust strategy for the multi-component analysis of food additives, thereby expanding the horizons of nanozymes in food safety monitoring.</div></div>","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"1413 ","pages":"Article 345668"},"PeriodicalIF":6.0,"publicationDate":"2026-05-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147999618","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}
Pub Date : 2026-04-15Epub Date: 2026-02-11DOI: 10.1016/j.aca.2026.345232
Nan Yang , Le Kang , Wei Yao , Baotong Xu , Yunhui Feng , Vladimir P. Fedin , Enjun Gao
Background
The global consumption of antibiotics is steadily rising, with doxycycline being extensively utilized in both animal husbandry and the treatment of COVID-19. Overuse of this antibiotic may result in residual accumulation and pose potential health risks. Conventional detection methods are often complex and labor-intensive, whereas fluorescence analysis offers advantages such as high sensitivity and rapid response. In this work, the development of a simple and efficient MOF-based fluorescent sensor for the detection of doxycycline holds considerable promise for practical applications.
Results
In this paper, a unique 3D metal-organic framework Zn[C18H16O4N4]·C2H6SO·2H2O (Zn-MOF) for rapid detection of doxycycline was successfully synthesized and analyzed by SEM, PXRD, FTIR, TGA and UV-vis. The fluorescence of Zn-MOF at 430 nm is quenched by the static quenching and inner filter effect between Zn-MOF and doxycycline with detection limit of 2.01 μM. Concurrently, Ca2+ can combine with doxycycline and form the Zn-MOF that generates a new fluorescence signal at 550 nm. By mixing the Zn-MOF and Ca2+, a ratiometric fluorescent probe can be created. The ratio of fluorescence intensity (F550/F430) to doxycycline concentration revealed a good linear relationship and detection limit of 1.25 μM. The fluorescence detection efficiency of the ratiometric fluorescent probe is almost twice that of the single metal-organic framework.
Significance
The ratiometric fluorescent probe also exhibited excellent selectivity and stability in both ionic and antibiotic solution. As with the concentration of doxycycline increase, the color of the probe and Ca2+ shifted from blue to yellow, allowing for visual detection of doxycycline and facilitating its application in rapid detection in real life.
{"title":"A ratiometric fluorescent probe consisting of Zn-MOF and calcium ion for sensitive and visual detection of doxycycline","authors":"Nan Yang , Le Kang , Wei Yao , Baotong Xu , Yunhui Feng , Vladimir P. Fedin , Enjun Gao","doi":"10.1016/j.aca.2026.345232","DOIUrl":"10.1016/j.aca.2026.345232","url":null,"abstract":"<div><h3>Background</h3><div>The global consumption of antibiotics is steadily rising, with doxycycline being extensively utilized in both animal husbandry and the treatment of COVID-19. Overuse of this antibiotic may result in residual accumulation and pose potential health risks. Conventional detection methods are often complex and labor-intensive, whereas fluorescence analysis offers advantages such as high sensitivity and rapid response. In this work, the development of a simple and efficient MOF-based fluorescent sensor for the detection of doxycycline holds considerable promise for practical applications.</div></div><div><h3>Results</h3><div>In this paper, a unique 3D metal-organic framework Zn[C<sub>18</sub>H<sub>16</sub>O<sub>4</sub>N<sub>4</sub>]·C<sub>2</sub>H<sub>6</sub>SO·2H<sub>2</sub>O (Zn-MOF) for rapid detection of doxycycline was successfully synthesized and analyzed by SEM, PXRD, FTIR, TGA and UV-vis. The fluorescence of Zn-MOF at 430 nm is quenched by the static quenching and inner filter effect between Zn-MOF and doxycycline with detection limit of 2.01 μM. Concurrently, Ca<sup>2+</sup> can combine with doxycycline and form the Zn-MOF that generates a new fluorescence signal at 550 nm. By mixing the Zn-MOF and Ca<sup>2+</sup>, a ratiometric fluorescent probe can be created. The ratio of fluorescence intensity (F<sub>550</sub>/F<sub>430</sub>) to doxycycline concentration revealed a good linear relationship and detection limit of 1.25 μM. The fluorescence detection efficiency of the ratiometric fluorescent probe is almost twice that of the single metal-organic framework.</div></div><div><h3>Significance</h3><div>The ratiometric fluorescent probe also exhibited excellent selectivity and stability in both ionic and antibiotic solution. As with the concentration of doxycycline increase, the color of the probe and Ca<sup>2+</sup> shifted from blue to yellow, allowing for visual detection of doxycycline and facilitating its application in rapid detection in real life.</div></div>","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"1395 ","pages":"Article 345232"},"PeriodicalIF":6.0,"publicationDate":"2026-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146187536","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}
Pub Date : 2026-04-15Epub Date: 2026-02-11DOI: 10.1016/j.aca.2026.345223
Haowei Ma , Ming Bi , Lang Zhao , Zhixin Tian
Background
Mass spectrometry-based shotgun proteomics relies on efficient sample preparation, where proteins are reduced, alkylated, and digested into peptides for LC-MS/MS analysis. While LC-MS/MS and bioinformatic identification have advanced to minute-scale workflows, sample preparation remains a major bottleneck, typically requiring hours to days. Although methods such as high-pressure and droplet-based reactions have reduced processing times, achieving minute-scale preparation has remained challenging. There is a clear need for an integrated, rapid sample preparation strategy to match the speed of modern LC-MS/MS and data analysis platforms.
Results
We developed OPPRAD (One-Pot Protein Reduction, Alkylation, and Digestion), an ultrafast sample preparation method conducted in water-in-oil nano/micro-droplets. This one-pot process completes all three key reactions within 5 min. When integrated with rapid LC-MS/MS on the Orbitrap Astral platform and MSFragger- or DIA–NN–based identification, the entire workflow achieved serum-to-peptide identification in 25.9 min and tissue-to-identification in 35.9 min—the fastest reported proteomic pipeline. The method showed high peptide recovery (∼68–71%), excellent cysteine alkylation (>99%), and miss cleavage rates comparable to conventional bulk digestion. It enabled deep proteome coverage across a wide dynamic range and was successfully applied to identify differentially expressed proteins in paired liver cancer tissues.
Significance
OPPRAD drastically shortens proteomic sample preparation from hours to minutes, reducing steps, cost, and hands-on time. This innovation bridges a critical gap in high-throughput proteomics, enabling rapid translational applications and large-scale clinical studies. The method's speed and efficiency pave the way for real-time proteomic analysis and future integration with online desalting and database searching.
基于质谱的霰弹枪蛋白质组学依赖于高效的样品制备，其中蛋白质被还原，烷基化，消化成肽，用于LC-MS/MS分析。虽然LC-MS/MS和生物信息学鉴定已经发展到分钟级的工作流程，但样品制备仍然是一个主要瓶颈，通常需要数小时到数天。尽管高压和液滴反应等方法减少了处理时间，但实现分钟级制备仍然具有挑战性。显然需要一个集成的，快速的样品制备策略，以匹配现代LC-MS/MS和数据分析平台的速度。结果建立了一种在油包水纳米/微滴中进行超快速样品制备的OPPRAD （One-Pot Protein Reduction, alkyation, and Digestion）方法。这一锅过程在5分钟内完成所有三个关键反应。当与Orbitrap Astral平台上的快速LC-MS/MS和基于MSFragger或dia - nn的鉴定相结合时，整个工作流程在25.9分钟内完成了血清到肽的鉴定，在35.9分钟内完成了组织到蛋白质组学的鉴定，这是目前报道的最快的蛋白质组学流程。该方法显示出高肽回收率（~ 68-71%），优异的半胱氨酸烷基化（>99%）和与传统批量消化相当的未裂解率。它能够在广泛的动态范围内实现深度蛋白质组覆盖，并成功地应用于鉴定配对肝癌组织中的差异表达蛋白。意义：opprad大大缩短了蛋白质组学样品制备从几小时到几分钟，减少了步骤，成本和动手时间。这一创新弥补了高通量蛋白质组学的关键空白，使快速转化应用和大规模临床研究成为可能。该方法的速度和效率为实时蛋白质组学分析和未来与在线脱盐和数据库搜索的集成铺平了道路。
{"title":"Ultrafast peptide preparation brings shotgun proteomics into the minute era","authors":"Haowei Ma , Ming Bi , Lang Zhao , Zhixin Tian","doi":"10.1016/j.aca.2026.345223","DOIUrl":"10.1016/j.aca.2026.345223","url":null,"abstract":"<div><h3>Background</h3><div>Mass spectrometry-based shotgun proteomics relies on efficient sample preparation, where proteins are reduced, alkylated, and digested into peptides for LC-MS/MS analysis. While LC-MS/MS and bioinformatic identification have advanced to minute-scale workflows, sample preparation remains a major bottleneck, typically requiring hours to days. Although methods such as high-pressure and droplet-based reactions have reduced processing times, achieving minute-scale preparation has remained challenging. There is a clear need for an integrated, rapid sample preparation strategy to match the speed of modern LC-MS/MS and data analysis platforms.</div></div><div><h3>Results</h3><div>We developed OPPRAD (One-Pot Protein Reduction, Alkylation, and Digestion), an ultrafast sample preparation method conducted in water-in-oil nano/micro-droplets. This one-pot process completes all three key reactions within 5 min. When integrated with rapid LC-MS/MS on the Orbitrap Astral platform and MSFragger- or DIA–NN–based identification, the entire workflow achieved serum-to-peptide identification in 25.9 min and tissue-to-identification in 35.9 min—the fastest reported proteomic pipeline. The method showed high peptide recovery (∼68–71%), excellent cysteine alkylation (>99%), and miss cleavage rates comparable to conventional bulk digestion. It enabled deep proteome coverage across a wide dynamic range and was successfully applied to identify differentially expressed proteins in paired liver cancer tissues.</div></div><div><h3>Significance</h3><div>OPPRAD drastically shortens proteomic sample preparation from hours to minutes, reducing steps, cost, and hands-on time. This innovation bridges a critical gap in high-throughput proteomics, enabling rapid translational applications and large-scale clinical studies. The method's speed and efficiency pave the way for real-time proteomic analysis and future integration with online desalting and database searching.</div></div>","PeriodicalId":240,"journal":{"name":"Analytica Chimica Acta","volume":"1395 ","pages":"Article 345223"},"PeriodicalIF":6.0,"publicationDate":"2026-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146161816","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}
Pub Date : 2026-04-15Epub Date: 2026-02-09DOI: 10.1016/j.aca.2026.345221
Guangxin Zhang , Haixu Zhao , Xiaoqi Li , Yi Zhang, Haijing Jia, Qiaoqiao Zhang, Tianyu Dai, Xinhui Lou
Background
The conflicted affinity evaluation results of small molecule-binding aptamers (SM-Apts) are increasing, raising the concerns on if a reported aptamer is a real aptamer. AMP17 is the first published controversial aptamer, which has been applied in at least 37 reported aptasensors for the detection of ampicillin, but was disproved by several homogeneous affinity methods including isothermal titration calorimetry, nuclear magnetic resonance spectroscopy, and mass spectrometry. Only the original paper reported its dissociation constant (KD = 13.4 nM) by the affinity column-based fluorescence method. If is AMP17 a real aptamer? What are the possible reasons that result in the conflicted results?
Results
We carried out four most popularly used homogeneous assays (FRET-based strand displacement assays, circular dichroism, EvaGreen and thioflavin T–based label-free fluorescence assays, and DNase I assay). None of them proved the binding of AMP17 to ampicillin. We then successfully measured the KD using a fiber optic evanescent wave sensor (FOEW), in which AMP17 was covalent immobilized on the SiO2 fiber without (KD = 2.6 ± 0.2 nM) or with BSA coating (KD = 5.1 ± 0.5 nM). The high specificity of AMP17 was proved by testing other molecules and replacing AMP17 by a non-binding sequence. We finally constructed a FOEW for the detection of ampicillin via one-step competitive or two step-noncompetitive detection modes, respectively. The latter showed 25.9-fold lower limit of detection (LOD = 2.8 fM, S/N = 3) than the former. Both detection modes possessed the good anti-matrix interference capability and were capable for the detection of ampicillin spiked in river water and milk without the need of sample purification.
Significance
Our results support that AMP17 is a real aptamer with high affinity and specificity. The surface immobilization favors the binding of AMP17 with ampicillin. Our study highlights the limited application scope of homogeneous assays for affinity evaluation of small molecule-binding aptamers. FOEW is a facile platform for both KD measurements and rapid detection.
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