Pub Date : 2025-02-21DOI: 10.1021/acs.analchem.4c06670
Peilin Wang, Zihui Liang, Wenyan Li, Zhenrun Li, Qiang Ma
The protein glycosylation of extracellular vesicles (EVs) is involved in cellular recognition and emerges as a promising biomarker for cancer diagnosis. However, the lack of efficient labeling and high-resolution detection strategies limits their clinical application. Herein, we developed a self-localized plasmonic nanocavity strategy to analyze the glycosylation characteristics of glioblastoma EVs. First, an engineered phospholipid bilayer structure with a Au nanoring array was designed to capture EVs and induce membrane fusion. Relying on the multifunctional proximity labeling process, a peroxidase-induced proximity labeling was designed to label sialic acid on programmed cell death ligand 1 (PD-L1) of EVs. Based on the identification and labeling process of EVs, the plasmonic nanocavity was self-localized with Au nanocubes and achieved the spontaneous location of MoSe2 QDs. The uniformly enhanced electromagnetic field in the nanocavity resulted in the polarized luminescence signal of MoSe2 QDs for improving the detection sensitivity and resolution. This system demonstrated the precise distinction and sensitive quantification of EV glycosylation in cerebrospinal fluid to distinguish glioblastoma. This research provided a novel strategy for the glycosylation detection of EVs and promoted the clinical application of EVs in glioblastoma diagnosis.
{"title":"Self-Localized Plasmonic Nanocavity Strategy for the Glycosylation Detection of Glioblastoma Extracellular Vesicles","authors":"Peilin Wang, Zihui Liang, Wenyan Li, Zhenrun Li, Qiang Ma","doi":"10.1021/acs.analchem.4c06670","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c06670","url":null,"abstract":"The protein glycosylation of extracellular vesicles (EVs) is involved in cellular recognition and emerges as a promising biomarker for cancer diagnosis. However, the lack of efficient labeling and high-resolution detection strategies limits their clinical application. Herein, we developed a self-localized plasmonic nanocavity strategy to analyze the glycosylation characteristics of glioblastoma EVs. First, an engineered phospholipid bilayer structure with a Au nanoring array was designed to capture EVs and induce membrane fusion. Relying on the multifunctional proximity labeling process, a peroxidase-induced proximity labeling was designed to label sialic acid on programmed cell death ligand 1 (PD-L1) of EVs. Based on the identification and labeling process of EVs, the plasmonic nanocavity was self-localized with Au nanocubes and achieved the spontaneous location of MoSe<sub>2</sub> QDs. The uniformly enhanced electromagnetic field in the nanocavity resulted in the polarized luminescence signal of MoSe<sub>2</sub> QDs for improving the detection sensitivity and resolution. This system demonstrated the precise distinction and sensitive quantification of EV glycosylation in cerebrospinal fluid to distinguish glioblastoma. This research provided a novel strategy for the glycosylation detection of EVs and promoted the clinical application of EVs in glioblastoma diagnosis.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"5 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143462162","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-21DOI: 10.1021/acs.analchem.4c05590
Iman Tahmasbian, Armando Navas, Mark W. Dunlop
Methane (CH4) is a significant greenhouse gas, and accurately quantifying its concentrations is essential for addressing climate change concerns. This study used controlled conditions to identify potential spectral regions or wavelengths within the short-wave infrared (SWIR) region that can be used for CH4 quantification using hyperspectral imaging (HSI). This study also validated the efficiency of the wavelengths that are currently used in remote sensing. Glass containers with constant CH4 flow rates were used for collecting HSI data (1010–2495 nm) at different CH4 concentrations (N = 18; 0–2.5% CH4). Partial least-squares regression (PLSR) was trained using the full 266-bands (1010–2495 nm). Regression coefficients and PLS weights were used to identify the potentially important regions and wavelengths. New PLSR models were developed using the important regions (multiband models). Individual wavelengths identified in the current study or previously used in remote sensing studies were also used to develop PLSR models individually or in two-band combinations. Potential overlaps between the identified spectral region and H2O absorption bands were investigated. The results indicated that using spectral regions (multiband) or combinations of two bands provided more accuracy compared to when single bands were used. The following spectral regions can be used for the quantification of CH4 in descending order: full 266-band (1010–2495 nm) > 1648 + 1670 nm > full 128-band (1010–1700 nm) > 2150–2243 nm > 1010–1185 nm. The results of this study were obtained under controlled conditions without interfering compounds. Testing these spectral regions in more complex environments will help confirm the best SWIR wavelengths.
{"title":"Feature Wavelengths for Quantifying Methane Concentrations Using Shortwave Infrared Hyperspectral Imaging: A Controlled Condition Study","authors":"Iman Tahmasbian, Armando Navas, Mark W. Dunlop","doi":"10.1021/acs.analchem.4c05590","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c05590","url":null,"abstract":"Methane (CH<sub>4</sub>) is a significant greenhouse gas, and accurately quantifying its concentrations is essential for addressing climate change concerns. This study used controlled conditions to identify potential spectral regions or wavelengths within the short-wave infrared (SWIR) region that can be used for CH<sub>4</sub> quantification using hyperspectral imaging (HSI). This study also validated the efficiency of the wavelengths that are currently used in remote sensing. Glass containers with constant CH<sub>4</sub> flow rates were used for collecting HSI data (1010–2495 nm) at different CH<sub>4</sub> concentrations (<i>N</i> = 18; 0–2.5% CH<sub>4</sub>). Partial least-squares regression (PLSR) was trained using the full 266-bands (1010–2495 nm). Regression coefficients and PLS weights were used to identify the potentially important regions and wavelengths. New PLSR models were developed using the important regions (multiband models). Individual wavelengths identified in the current study or previously used in remote sensing studies were also used to develop PLSR models individually or in two-band combinations. Potential overlaps between the identified spectral region and H<sub>2</sub>O absorption bands were investigated. The results indicated that using spectral regions (multiband) or combinations of two bands provided more accuracy compared to when single bands were used. The following spectral regions can be used for the quantification of CH<sub>4</sub> in descending order: full 266-band (1010–2495 nm) > 1648 + 1670 nm > full 128-band (1010–1700 nm) > 2150–2243 nm > 1010–1185 nm. The results of this study were obtained under controlled conditions without interfering compounds. Testing these spectral regions in more complex environments will help confirm the best SWIR wavelengths.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"15 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143462223","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-21DOI: 10.1021/acs.analchem.4c03985
Johnnie Phuong, Billy Salgado, Tom Labusch, Hans Hasse, Kerstin Münnemann
Benchtop 13C NMR spectroscopy is highly attractive for reaction and process monitoring. However, insufficient premagnetization and low signal intensities largely prevent its application to flowing liquids. We show that hyperpolarization by Overhauser dynamic nuclear polarization (ODNP) can be used to overcome these problems, as ODNP operates on short time scales and results in strong 13C signal enhancements. Benchtop 13C NMR spectra with ODNP enhancement acquired in continuous-flow are reported here for the first time. We have investigated two ODNP approaches: direct ODNP, which transfers the polarization of unpaired electrons to 13C nuclei via direct hyperfine coupling, and indirect ODNP, in which the electron polarization is first transferred to 1H nuclei before a polarization transfer pulse sequence finally transfers the polarization to the 13C nuclei. Experiments were carried out for three pure solvents and a mixture for different flow rates. The results show significant 13C signal enhancements for both approaches. However, their performance varies for different substances, depending on the strength and type of the hyperfine interaction as well as on the relaxation properties, but by selecting a suitable approach, good single-scan 13C NMR spectra can be obtained with benchtop NMR, even at high flow rates.
{"title":"Overhauser Dynamic Nuclear Polarization Enables Single Scan Benchtop 13C NMR Spectroscopy in Continuous-Flow","authors":"Johnnie Phuong, Billy Salgado, Tom Labusch, Hans Hasse, Kerstin Münnemann","doi":"10.1021/acs.analchem.4c03985","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c03985","url":null,"abstract":"Benchtop <sup>13</sup>C NMR spectroscopy is highly attractive for reaction and process monitoring. However, insufficient premagnetization and low signal intensities largely prevent its application to flowing liquids. We show that hyperpolarization by Overhauser dynamic nuclear polarization (ODNP) can be used to overcome these problems, as ODNP operates on short time scales and results in strong <sup>13</sup>C signal enhancements. Benchtop <sup>13</sup>C NMR spectra with ODNP enhancement acquired in continuous-flow are reported here for the first time. We have investigated two ODNP approaches: direct ODNP, which transfers the polarization of unpaired electrons to <sup>13</sup>C nuclei via direct hyperfine coupling, and indirect ODNP, in which the electron polarization is first transferred to <sup>1</sup>H nuclei before a polarization transfer pulse sequence finally transfers the polarization to the <sup>13</sup>C nuclei. Experiments were carried out for three pure solvents and a mixture for different flow rates. The results show significant <sup>13</sup>C signal enhancements for both approaches. However, their performance varies for different substances, depending on the strength and type of the hyperfine interaction as well as on the relaxation properties, but by selecting a suitable approach, good single-scan <sup>13</sup>C NMR spectra can be obtained with benchtop NMR, even at high flow rates.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"89 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143470897","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
In the current study, we designed a unique core-to-shell thickness-regulated Ag@Au nanocatalyst (CSNPs) for H2O2-induced selective oxidative etching of core silver. Synthesized CSNPs exhibit high colloidal stability and demonstrate a significant localized surface plasmon resonance (LSPR) effect in the biological window. These unique properties in turn allow us to formulate a unique CSNP-based LSPR-induced electrochemical detection assay for selective trace-level sensing of H2O2in vitro. Conceptually, we utilized LSPR to amplify the electrochemical signals by inducing the generation of hot electrons and hot holes, which can be harnessed for catalytic purposes. Here, the Au shell acts as a supplier of the hot electron for enhanced catalytic reduction of H2O2 where the free electron of the Au shell is subsidized by the Ag core by its subsequent oxidation. The combination of high LSPR property, stability, and efficient binding property makes these NPs not only a surface-enhanced Raman scattering (SERS) enhancer but also a promising electrocatalyst for biomolecule detection, which emphasizes the significant potential of these engineered nanomaterials in various applications.
{"title":"Core-to-Shell Thickness-Regulated Ag@Au Nanocatalyst for LSPR-Improved In Situ Detection of Extracellular Peroxide: Response in a Cancer Cell","authors":"Manorama Ghosal, Subrata Mondal, Tanmay Ghosh, Debasish Prusty, Dulal Senapati","doi":"10.1021/acs.analchem.4c04651","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c04651","url":null,"abstract":"In the current study, we designed a unique core-to-shell thickness-regulated Ag@Au nanocatalyst (<sub>CS</sub>NPs) for H<sub>2</sub>O<sub>2</sub>-induced selective oxidative etching of core silver. Synthesized <sub>CS</sub>NPs exhibit high colloidal stability and demonstrate a significant localized surface plasmon resonance (LSPR) effect in the biological window. These unique properties in turn allow us to formulate a unique <sub>CS</sub>NP-based LSPR-induced electrochemical detection assay for selective trace-level sensing of H<sub>2</sub>O<sub>2</sub> <i>in vitro</i>. Conceptually, we utilized LSPR to amplify the electrochemical signals by inducing the generation of hot electrons and hot holes, which can be harnessed for catalytic purposes. Here, the Au shell acts as a supplier of the hot electron for enhanced catalytic reduction of H<sub>2</sub>O<sub>2</sub> where the free electron of the Au shell is subsidized by the Ag core by its subsequent oxidation. The combination of high LSPR property, stability, and efficient binding property makes these NPs not only a surface-enhanced Raman scattering (SERS) enhancer but also a promising electrocatalyst for biomolecule detection, which emphasizes the significant potential of these engineered nanomaterials in various applications.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"5 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143470898","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Microplastics (MPs) have been universally recognized as a pervasive and enduring environmental hazard, promoting research on relevant analytical techniques. Despite the unique advantages of mass spectrometry (MS) for polymer identification, lengthy procedures and complex data processing are always ineluctable. In this study, an ambient microwave plasma torch (MPT) ion source coupled with an LTQ Orbitrap MS was developed, presenting a rapid and concise analytical approach for MPs with simplified pretreatment and intuitive mass spectra. One testing process took approximately 30 s, enabling a higher efficiency of analysis. Furthermore, the method was not constrained by the MP size limitation; even macroscale polymer blocks could be detected. Under the optimized conditions, the method was proven to be efficient for the desorption and ionization of a wide range of MPs (polyamide, poly(ethylene terephthalate), polymethacrylate, polylactic acid, poly(3-hydroxybutyrate), polypropylene, and polythene), while the distinctly decipherable spectra intuitively reflected the mass intervals conforming to the corresponding monomer of MPs. Linear relationships were established between sample mass and the intensity of characteristic ion, with R2 exceeding 0.98. Additionally, a simplified pretreatment process in conjunction with MPT-MS was explored, verifying the method’s resilience to matrix interferences and its applicability to environmental sample analysis. Furthermore, the compatibility of the established method with scanning electron microscopy was taken into consideration, thereby complementing traditional MS analysis by providing additional insights into the size and morphology of MPs. This study employed MPT as the ion source for MS detection of MPs, establishing a concise, rapid, and comprehensive method specifically targeting the analysis of MPs, which provided inspiration for the extraction and characterization of MPs in environmental samples.
{"title":"Concise, Rapid, and Comprehensive Approach for Microplastic Detection Based on Ambient Microwave Plasma Torch Desorption/Ionization Mass Spectrometry","authors":"Qing Li, Weiwei Chen, Fengjian Chu, Jing Luo, Hongru Feng, Yuanjiang Pan","doi":"10.1021/acs.analchem.4c05789","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c05789","url":null,"abstract":"Microplastics (MPs) have been universally recognized as a pervasive and enduring environmental hazard, promoting research on relevant analytical techniques. Despite the unique advantages of mass spectrometry (MS) for polymer identification, lengthy procedures and complex data processing are always ineluctable. In this study, an ambient microwave plasma torch (MPT) ion source coupled with an LTQ Orbitrap MS was developed, presenting a rapid and concise analytical approach for MPs with simplified pretreatment and intuitive mass spectra. One testing process took approximately 30 s, enabling a higher efficiency of analysis. Furthermore, the method was not constrained by the MP size limitation; even macroscale polymer blocks could be detected. Under the optimized conditions, the method was proven to be efficient for the desorption and ionization of a wide range of MPs (polyamide, poly(ethylene terephthalate), polymethacrylate, polylactic acid, poly(3-hydroxybutyrate), polypropylene, and polythene), while the distinctly decipherable spectra intuitively reflected the mass intervals conforming to the corresponding monomer of MPs. Linear relationships were established between sample mass and the intensity of characteristic ion, with <i>R</i><sup>2</sup> exceeding 0.98. Additionally, a simplified pretreatment process in conjunction with MPT-MS was explored, verifying the method’s resilience to matrix interferences and its applicability to environmental sample analysis. Furthermore, the compatibility of the established method with scanning electron microscopy was taken into consideration, thereby complementing traditional MS analysis by providing additional insights into the size and morphology of MPs. This study employed MPT as the ion source for MS detection of MPs, establishing a concise, rapid, and comprehensive method specifically targeting the analysis of MPs, which provided inspiration for the extraction and characterization of MPs in environmental samples.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"67 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143470920","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1021/acs.analchem.4c05961
Yan Li, Yichen Li, Yueli Hu, Rui Liu, Yi Lv
The CRISPR-Cas system, particularly CRISPR-Cas12a and CRISPR-Cas13a, has been widely utilized in constructing various biosensors due to their “trans-cleavage” ability as a means of signal amplification. However, this universal “trans-cleavage” characteristic also presents a challenge for realizing CRISPR-Cas multiplexed bioanalysis. Besides, potential signal cascading interference and complicated design are notable obstacles in CRISPR-Cas multiplexed bioanalysis. Herein, we propose a mass spectrometry method that leverages the CRISPR-Cas12a/13a system to achieve simultaneous detection of ctDNA and miRNA. Based on the properties of the CRISPR-Cas12a/13a system, two types of nanoparticle reporter probes have been engineered, using cancer-related biomarkers ctDNA and miR-21 as our model analytes. The nanoparticle tags, which intrinsically incorporated millions of detectable atoms, combined with the CRISPR-Cas12a/Cas13a system’s “trans-cleavage” ability, allow the proposed mass spectrometry strategy to achieve fmol-level detection limits without any nucleic acid amplification procedures. The assay was successfully applied to human serum samples, demonstrating its potential for early disease diagnosis and progression tracking.
{"title":"CRISPR-Cas12a/Cas13a Multiplex Bioassay for ctDNA and miRNA by Mass Spectrometry","authors":"Yan Li, Yichen Li, Yueli Hu, Rui Liu, Yi Lv","doi":"10.1021/acs.analchem.4c05961","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c05961","url":null,"abstract":"The CRISPR-Cas system, particularly CRISPR-Cas12a and CRISPR-Cas13a, has been widely utilized in constructing various biosensors due to their “<i>trans</i>-cleavage” ability as a means of signal amplification. However, this universal “<i>trans</i>-cleavage” characteristic also presents a challenge for realizing CRISPR-Cas multiplexed bioanalysis. Besides, potential signal cascading interference and complicated design are notable obstacles in CRISPR-Cas multiplexed bioanalysis. Herein, we propose a mass spectrometry method that leverages the CRISPR-Cas12a/13a system to achieve simultaneous detection of ctDNA and miRNA. Based on the properties of the CRISPR-Cas12a/13a system, two types of nanoparticle reporter probes have been engineered, using cancer-related biomarkers ctDNA and miR-21 as our model analytes. The nanoparticle tags, which intrinsically incorporated millions of detectable atoms, combined with the CRISPR-Cas12a/Cas13a system’s “<i>trans</i>-cleavage” ability, allow the proposed mass spectrometry strategy to achieve fmol-level detection limits without any nucleic acid amplification procedures. The assay was successfully applied to human serum samples, demonstrating its potential for early disease diagnosis and progression tracking.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"27 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143462229","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1021/acs.analchem.4c05037
Julia Järlebark, Wei Liu, Amina Shaji, Jingjie Sha, Andreas Dahlin
Solid state nanopores have emerged as an important electrical label-free single-molecule detection platform. While much effort has been spent on analyzing the current trace to determine size, shape and charge of the translocating species, a less studied aspect is the number of events and how this relates to analyte concentration. In this work we systematically investigate how the event frequency depends on voltage applied across the pore and show that this dependence can be utilized to determine target concentration. Importantly, this method does not require any calibration or any additional species added to the solution. Data analysis algorithms are introduced to accurately count events also for high voltages (up to 1 V). For double stranded DNA as model analyte, we find a linear relation between event frequency and voltage for pores 10 nm or more in diameter. For smaller pores, the majority of events are dockings rather than translocations and the linear relation is lost, in agreement with theory. Our model also predicts that the electrophoretic mobility of the species will influence event frequency, while diffusivity does not, which we confirm by using two different sizes of DNA. The analyte concentration determination is found to be remarkably accurate (10% error) when taking the average of multiple (∼4) experiments. If based on a single experiment, the predictive power is lower, but the method still provides a useful estimate (<30% error). This study should be useful as a guide when performing experiments at higher voltages and may serve as a method to extract analyte concentration in bioanalytical applications of nanopore sensors.
固态纳米孔已经成为一种重要的无电学标记单分子检测平台。虽然人们已经在分析电流轨迹以确定转运物种的大小、形状和电荷方面花费了大量精力，但对事件发生的次数及其与分析物浓度的关系研究较少。在这项工作中，我们系统地研究了事件频率如何依赖于施加在孔隙上的电压，并证明可以利用这种依赖关系来确定目标浓度。重要的是，这种方法不需要任何校准，也不需要向溶液中添加任何其他物质。介绍了数据分析算法，以便在高电压（高达 1 V）条件下也能对事件进行精确计数。对于以双链 DNA 为模型的分析物，我们发现对于直径为 10 nm 或更大的孔，事件频率与电压之间呈线性关系。对于更小的孔，大多数事件是对接而不是转移，因此线性关系消失，这与理论一致。我们的模型还预测，物种的电泳迁移率会影响事件频率，而扩散率不会，我们使用两种不同大小的 DNA 证实了这一点。当取多个（∼4）实验的平均值时，我们发现分析物浓度的测定非常准确（误差为 10%）。如果基于单次实验，预测能力较低，但该方法仍能提供有用的估计值（误差 30%）。这项研究对在更高电压下进行实验具有指导作用，并可作为纳米孔传感器生物分析应用中提取分析物浓度的一种方法。
{"title":"Solid-State Nanopore Sensors: Analyte Quantification by Event Frequency Analysis at High Voltages","authors":"Julia Järlebark, Wei Liu, Amina Shaji, Jingjie Sha, Andreas Dahlin","doi":"10.1021/acs.analchem.4c05037","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c05037","url":null,"abstract":"Solid state nanopores have emerged as an important electrical label-free single-molecule detection platform. While much effort has been spent on analyzing the current trace to determine size, shape and charge of the translocating species, a less studied aspect is the number of events and how this relates to analyte concentration. In this work we systematically investigate how the event frequency depends on voltage applied across the pore and show that this dependence can be utilized to determine target concentration. Importantly, this method does not require any calibration or any additional species added to the solution. Data analysis algorithms are introduced to accurately count events also for high voltages (up to 1 V). For double stranded DNA as model analyte, we find a linear relation between event frequency and voltage for pores 10 nm or more in diameter. For smaller pores, the majority of events are dockings rather than translocations and the linear relation is lost, in agreement with theory. Our model also predicts that the electrophoretic mobility of the species will influence event frequency, while diffusivity does not, which we confirm by using two different sizes of DNA. The analyte concentration determination is found to be remarkably accurate (10% error) when taking the average of multiple (∼4) experiments. If based on a single experiment, the predictive power is lower, but the method still provides a useful estimate (<30% error). This study should be useful as a guide when performing experiments at higher voltages and may serve as a method to extract analyte concentration in bioanalytical applications of nanopore sensors.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"85 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143452149","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1021/acs.analchem.4c06146
Yunshan Zhang, Tuo Huang, Fang Yang, Qianglong Tan, Jing Ye, Xianzhong Feng, Diming Zhang
In recent years, entropy-driven circuit (EDC) dynamic DNA networks have garnered significant attention in nucleic acid detection owing to their simplicity, efficiency, and flexible design. Nevertheless, conventional EDC reactions face a constraint in achieving optimal signal amplification due to a solitary and feeble driving force. To overcome this limitation, we innovatively devised a gold nanoparticle (AuNP) dispersion-enhanced EDC (Au-EDC) approach, pioneering a novel colorimetric signal amplification and output system. The system was harmoniously integrated with the ligase chain reaction (LCR) for precise single nucleotide polymorphism (SNP) genotyping. Specifically, LCR was selectively executed solely on the positive strand of the mutant target (MT), facilitating precise point-to-strand information transduction. Subsequently, the LCR product triggered the Au-EDC cycling reaction, causing the DNA-AuNPs network to progressively disintegrate and release a pronounced colorimetric signal. This strategic design ingeniously harnessed the entropy increase that occurs as AuNPs undergo a transition from aggregated to dispersed states, offering a supplemental impetus for the EDC cycle. The integrated LCR-Au-EDC system excelled in detecting MT at concentrations as low as 320 fM and differentiating pooled samples with mutation frequencies as low as 0.1%. Moreover, the system accurately performed SNP genotyping on the real genomes derived from soybean leaves. Consequently, this study not only develops a colorimetric signal amplification and output sensing system based on EDC reactions but also provides a cost-effective and efficient SNP genotyping tool.
近年来，熵驱动电路（EDC）动态 DNA 网络因其简单、高效和设计灵活而在核酸检测领域备受关注。然而，传统的 EDC 反应由于驱动力单一且微弱，在实现最佳信号放大方面面临着限制。为了克服这一限制，我们创新性地设计了一种金纳米粒子（AuNP）分散增强 EDC（Au-EDC）方法，开创了一种新型比色信号放大和输出系统。该系统与连接酶链反应（LCR）和谐地结合在一起，用于精确的单核苷酸多态性（SNP）基因分型。具体来说，LCR 只在突变目标（MT）的阳性链上选择性地执行，从而促进了精确的点到链信息转导。随后，LCR 产物触发 Au-EDC 循环反应，使 DNA-AuNPs 网络逐渐解体并释放出明显的比色信号。这一战略性设计巧妙地利用了 AuNPs 从聚集状态过渡到分散状态时发生的熵增加，为 EDC 循环提供了补充动力。LCR-Au-EDC 集成系统在检测浓度低至 320 fM 的 MT 和区分突变频率低至 0.1% 的集合样本方面表现出色。此外，该系统还能对来自大豆叶片的真实基因组进行准确的 SNP 基因分型。因此，本研究不仅开发了一种基于 EDC 反应的比色信号放大和输出传感系统，还提供了一种经济高效的 SNP 基因分型工具。
{"title":"Entropy-Driven Circuit Integrated with Ligases to Regulate DNA-AuNP Network Disintegration for Colorimetric Detection of Single Nucleotide Polymorphisms","authors":"Yunshan Zhang, Tuo Huang, Fang Yang, Qianglong Tan, Jing Ye, Xianzhong Feng, Diming Zhang","doi":"10.1021/acs.analchem.4c06146","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c06146","url":null,"abstract":"In recent years, entropy-driven circuit (EDC) dynamic DNA networks have garnered significant attention in nucleic acid detection owing to their simplicity, efficiency, and flexible design. Nevertheless, conventional EDC reactions face a constraint in achieving optimal signal amplification due to a solitary and feeble driving force. To overcome this limitation, we innovatively devised a gold nanoparticle (AuNP) dispersion-enhanced EDC (Au-EDC) approach, pioneering a novel colorimetric signal amplification and output system. The system was harmoniously integrated with the ligase chain reaction (LCR) for precise single nucleotide polymorphism (SNP) genotyping. Specifically, LCR was selectively executed solely on the positive strand of the mutant target (MT), facilitating precise point-to-strand information transduction. Subsequently, the LCR product triggered the Au-EDC cycling reaction, causing the DNA-AuNPs network to progressively disintegrate and release a pronounced colorimetric signal. This strategic design ingeniously harnessed the entropy increase that occurs as AuNPs undergo a transition from aggregated to dispersed states, offering a supplemental impetus for the EDC cycle. The integrated LCR-Au-EDC system excelled in detecting MT at concentrations as low as 320 fM and differentiating pooled samples with mutation frequencies as low as 0.1%. Moreover, the system accurately performed SNP genotyping on the real genomes derived from soybean leaves. Consequently, this study not only develops a colorimetric signal amplification and output sensing system based on EDC reactions but also provides a cost-effective and efficient SNP genotyping tool.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"50 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143452197","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1021/acs.analchem.4c06453
Yuqi Zeng, Zichun Qiao, He Wang, Bo Gao, Qianqian Jiang, Yukui Zhang, Lihua Zhang, Bo Jiang
Photocatalytic proximity labeling techniques enable significant advances in understanding the subcellular proteome. However, current methods primarily utilize visible light that suffers from low labeling efficiency and limited identification coverage due to absorption overlap with endogenous chromophores and scattering in biological samples. To address these issues, we reported the proximity labeling strategy using near-infrared excitation (PL-NIR) strategy, a proximity labeling platform that used a near-infrared-excited catalyst to activate labeling mediated by reactive oxygen species, which could avoid the disadvantages of visible light. Taking advantage of the near-infrared excitation and mitochondrial targeting properties of IR780, the mitochondrial proteome were selectively labeled by spatially limited reaction in the native environment. The PL-NIR strategy facilitated the plotting of the mitochondrial proteome in which up to 245 mitochondrial proteins were identified in living HeLa cells. Compared with the current methods, the PL-NIR strategy significantly increased identification coverage, especially for mitochondrial inner membrane proteins. Furthermore, mitochondrial proteome dynamics were deciphered in LPS stimulated HMC3 cells, which were hard to transfect. Overall, the PL-NIR strategy as a highly precise proteomic platform offered improved identification coverage for more knowledge of subcellular biology discovering.
{"title":"Near-Infrared Light-Mediated Living Cells Mitochondrial Proteome Proximity Labeling","authors":"Yuqi Zeng, Zichun Qiao, He Wang, Bo Gao, Qianqian Jiang, Yukui Zhang, Lihua Zhang, Bo Jiang","doi":"10.1021/acs.analchem.4c06453","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c06453","url":null,"abstract":"Photocatalytic proximity labeling techniques enable significant advances in understanding the subcellular proteome. However, current methods primarily utilize visible light that suffers from low labeling efficiency and limited identification coverage due to absorption overlap with endogenous chromophores and scattering in biological samples. To address these issues, we reported the proximity labeling strategy using near-infrared excitation (PL-NIR) strategy, a proximity labeling platform that used a near-infrared-excited catalyst to activate labeling mediated by reactive oxygen species, which could avoid the disadvantages of visible light. Taking advantage of the near-infrared excitation and mitochondrial targeting properties of IR780, the mitochondrial proteome were selectively labeled by spatially limited reaction in the native environment. The PL-NIR strategy facilitated the plotting of the mitochondrial proteome in which up to 245 mitochondrial proteins were identified in living HeLa cells. Compared with the current methods, the PL-NIR strategy significantly increased identification coverage, especially for mitochondrial inner membrane proteins. Furthermore, mitochondrial proteome dynamics were deciphered in LPS stimulated HMC3 cells, which were hard to transfect. Overall, the PL-NIR strategy as a highly precise proteomic platform offered improved identification coverage for more knowledge of subcellular biology discovering.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"25 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143452152","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1021/acs.analchem.4c04283
Dario Valter Conca, Fouzia Bano, Małgorzata Graul, Julius von Wirén, Lauriane Scherrer, Hudson Pace, Himanshu Sharma, Justas Svirelis, Konrad Thorsteinsson, Andreas Dahlin, Marta Bally
The spread of SARS-CoV-2 led to the emergence of several variants of concern (VOCs). The spike glycoprotein, responsible for engaging the viral receptor, exhibits the highest density of mutations, suggesting an ongoing evolution to optimize viral entry. This study characterizes the bond formed by virion mimics carrying the SARS-CoV-2 spike protein and the plasma membrane of host cells in the early stages of virus entry. Contrary to the traditional analysis of isolated ligand–receptor pairs, we utilized well-defined biomimetic models and biochemical and biophysical techniques to characterize the multivalent interaction of VOCs with the complex cell membrane. We observed an overall increase in the binding affinity for newer VOCs. By progressively reducing the system complexity, we identify heparan sulfate (HS) as a main driver of this variation, with a 10-fold increase in affinity for Omicron BA.1 over that of the original strain. These results demonstrate the essential role of coreceptors, particularly HS, in the modulation of SARS-CoV-2 infection and highlight the importance of multiscale biophysical and biochemical assays that account for membrane complexity to fully characterize and understand the role of molecular components and their synergy in viral attachment and entry.
{"title":"Variant-Specific Interactions at the Plasma Membrane: Heparan Sulfate’s Impact on SARS-CoV-2 Binding Kinetics","authors":"Dario Valter Conca, Fouzia Bano, Małgorzata Graul, Julius von Wirén, Lauriane Scherrer, Hudson Pace, Himanshu Sharma, Justas Svirelis, Konrad Thorsteinsson, Andreas Dahlin, Marta Bally","doi":"10.1021/acs.analchem.4c04283","DOIUrl":"https://doi.org/10.1021/acs.analchem.4c04283","url":null,"abstract":"The spread of SARS-CoV-2 led to the emergence of several variants of concern (VOCs). The spike glycoprotein, responsible for engaging the viral receptor, exhibits the highest density of mutations, suggesting an ongoing evolution to optimize viral entry. This study characterizes the bond formed by virion mimics carrying the SARS-CoV-2 spike protein and the plasma membrane of host cells in the early stages of virus entry. Contrary to the traditional analysis of isolated ligand–receptor pairs, we utilized well-defined biomimetic models and biochemical and biophysical techniques to characterize the multivalent interaction of VOCs with the complex cell membrane. We observed an overall increase in the binding affinity for newer VOCs. By progressively reducing the system complexity, we identify heparan sulfate (HS) as a main driver of this variation, with a 10-fold increase in affinity for Omicron BA.1 over that of the original strain. These results demonstrate the essential role of coreceptors, particularly HS, in the modulation of SARS-CoV-2 infection and highlight the importance of multiscale biophysical and biochemical assays that account for membrane complexity to fully characterize and understand the role of molecular components and their synergy in viral attachment and entry.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"2 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143452125","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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