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Three-Dimensional DNA Hydrogel Mediated Dual-Mode Sensing Method for Quantification of Epithelial Cell Adhesion Molecule in Biological Fluid Samples. 三维 DNA 水凝胶介导的双模传感法,用于定量检测生物液体样本中的上皮细胞粘附分子。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-15 DOI: 10.1021/acs.analchem.4c01006
Lu Huang, Hanbing Huang, Zhuomin Zhang, Gongke Li

Monitoring changes in the expression of marker proteins in biological fluids is essential for biomarker-based disease diagnosis. Epithelial cell adhesion molecule (EpCAM) has been identified as a broad-spectrum biomarker for various chronic diseases and as a therapeutic target. However, the development of simple and reliable methods for quantifying EpCAM changes in biological fluids faces challenges due to the variability of its expression across different diseases, the presence of soluble forms, and matrix effects. In this paper, a surface-enhanced Raman scattering (SERS)-fluorescence (FL) dual-mode sensing method was established for quantification of trace EpCAM in biological fluids based on bimetallic Au@Ag nanoparticles and nitrogen-doped quantum dots encapsulated DNA hydrogel hybrid with graphene oxide (Au@Ag-NQDs/GO). The DNA hydrogel was constructed based on three-dimensional (3D) structure DNA-mediated strategy using an aptamer DNA (AptDNA) linker. The interaction of the AptDNA with EpCAM triggered the disassembly of the DNA hydrogel. Consequently, the release of Au@Ag nanoparticles induced an "on-off" switch in the SERS signal while the weakened FL quenching effect in Au@Ag-NQDs/GO system achieved "off-on" switch of FL signal, enabling the simultaneous SERS-FL quantification of EpCAM. The established dual-mode method exhibited outstanding sensitivity and stability in quantifying EpCAM in the range of 0.5-60.0 pg/mL, with the limits of detection (LODs) of SERS and FL as 0.17 and 0.35 pg/mL, respectively. When applied for real sample analysis, the method showed satisfactory specificity and recoveries in cancer cells lysate, serum, and urine samples with RSDs of 2.8-6.3%, 4.0-6.3%, and 2.8-5.7%, respectively. The developed SERS-FL sensing method offered a sensitive, reliable, and practical quantification strategy for trace EpCAM in diverse biological fluid samples, which would benefit the early diagnosis of disease and further health management.

监测生物液体中标记蛋白的表达变化对于基于生物标记的疾病诊断至关重要。上皮细胞粘附分子(EpCAM)已被确定为各种慢性疾病的广谱生物标记物和治疗靶点。然而,由于 EpCAM 在不同疾病中的表达差异、可溶性形式的存在以及基质效应,开发简单可靠的方法来量化 EpCAM 在生物液体中的变化面临着挑战。本文建立了一种表面增强拉曼散射(SERS)-荧光(FL)双模式传感方法,基于双金属Au@Ag纳米粒子和氮掺杂量子点包裹氧化石墨烯杂交DNA水凝胶(Au@Ag-NQDs/GO)来定量检测生物液体中的痕量EpCAM。DNA 水凝胶是基于三维(3D)结构的 DNA 介导策略,使用 Aptamer DNA(AptDNA)链接器构建的。AptDNA 与 EpCAM 的相互作用引发了 DNA 水凝胶的解体。因此,Au@Ag 纳米粒子的释放诱导了 SERS 信号的 "开关 "切换,而 Au@Ag-NQDs/GO 系统中的荧光淬灭效应减弱,实现了荧光信号的 "关闭 "切换,从而实现了 EpCAM 的 SERS-FL 同步定量。所建立的双模式方法在0.5-60.0 pg/mL范围内对EpCAM的定量具有出色的灵敏度和稳定性,SERS和FL的检出限(LOD)分别为0.17和0.35 pg/mL。在实际样品分析中,该方法在癌细胞裂解液、血清和尿液样品中显示出令人满意的特异性和回收率,RSD分别为2.8-6.3%、4.0-6.3%和2.8-5.7%。所开发的SERS-FL传感方法为多种生物液体样品中痕量EpCAM的检测提供了一种灵敏、可靠和实用的定量策略,这将有利于疾病的早期诊断和进一步的健康管理。
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引用次数: 0
Rapid Microbial Profiling through Multimodal Biosensors for Transversal Analysis. 通过多模态生物传感器进行横向分析,实现快速微生物轮廓分析。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-15 DOI: 10.1021/acs.analchem.4c02449
Jyong-Huei Lee, Siew Mei Chin, Dennis C Chan, Joseph C Liao, Samuel Yang, Nanying Zhang, Pak Kin Wong

The intricate interactions between host and microbial communities hold significant implications for biology and medicine. However, traditional microbial profiling methods face limitations in processing time, measurement of absolute abundance, detection of low biomass, discrimination between live and dead cells, and functional analysis. This study introduces a rapid multimodal microbial characterization platform, Multimodal Biosensors for Transversal Analysis (MBioTA), for capturing the taxonomy, viability, and functional genes of the microbiota. The platform incorporates single cell biosensors, scalable microwell arrays, and automated image processing for rapid transversal analysis in as few as 2 h. The multimodal biosensors simultaneously characterize the taxon, viability, and functional gene expression of individual cells. By automating the image processing workflow, the single cell analysis techniques enable the quantification of bacteria with sensitivity down to 0.0075%, showcasing its capability in detecting low biomass samples. We illustrate the applicability of the MBioTA platform through the transversal analysis of the gut microbiota composition, viability, and functionality in a familial Alzheimer's disease mouse model. The effectiveness, rapid turnaround, and scalability of the MBioTA platform will facilitate its application from basic research to clinical diagnostics, potentially revolutionizing our understanding and management of diseases associated with microbe-host interactions.

宿主与微生物群落之间错综复杂的相互作用对生物学和医学有着重要影响。然而,传统的微生物表征方法在处理时间、绝对丰度测量、低生物量检测、活细胞和死细胞区分以及功能分析等方面存在局限性。本研究介绍了一种快速多模态微生物表征平台--横向分析多模态生物传感器(MBioTA),用于捕捉微生物区系的分类、活力和功能基因。该平台集成了单细胞生物传感器、可扩展的微孔阵列和自动图像处理功能,可在短短 2 小时内完成快速横向分析。通过图像处理工作流程的自动化,单细胞分析技术能够对细菌进行定量分析,灵敏度低至 0.0075%,展示了其检测低生物量样品的能力。我们通过横向分析家族性阿尔茨海默病小鼠模型的肠道微生物群组成、活力和功能,说明了 MBioTA 平台的适用性。MBioTA 平台的有效性、快速周转性和可扩展性将促进其从基础研究到临床诊断的应用,有可能彻底改变我们对与微生物-宿主相互作用相关疾病的理解和管理。
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引用次数: 0
An Optimized Miniaturized Filter-Aided Sample Preparation Method for Sensitive Cross-Linking Mass Spectrometry Analysis of Microscale Samples. 用于微尺度样品灵敏交联质谱分析的优化微型过滤器辅助样品制备方法。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-15 DOI: 10.1021/acs.analchem.4c01600
Yu He, Yang Li, Lili Zhao, Guojin Ying, Gang Lu, Lihua Zhang, Zhenbin Zhang

Cross-linking mass spectrometry (XL-MS) is a powerful tool for elucidating protein structures and protein-protein interactions (PPIs) at the global scale. However, sensitive XL-MS analysis of mass-limited samples remains challenging, due to serious sample loss during sample preparation of the low-abundance cross-linked peptides. Herein, an optimized miniaturized filter-aided sample preparation (O-MICROFASP) method was presented for sensitive XL-MS analysis of microscale samples. By systematically investigating and optimizing crucial experimental factors, this approach dramatically improves the XL identification of low and submicrogram samples. Compared with the conventional FASP method, more than 7.4 times cross-linked peptides were identified from single-shot analysis of 1 μg DSS cross-linked HeLa cell lysates (440 vs 59). The number of cross-linked peptides identified from 0.5 μg HeLa cell lysates was increased by 58% when further reducing the surface area of the filter to 0.058 mm2 in the microreactor. To deepen the identification coverage of XL-proteome, five different types of cross-linkers were used and each μg of cross-linked HeLa cell lysates was processed by O-MICROFASP integrated with tip-based strong cation exchange (SCX) fractionation. Up to 2741 unique cross-linked peptides were identified from the 5 μg HeLa cell lysates, representing 2579 unique K-K linkages on 1092 proteins. About 96% of intraprotein cross-links were within the maximal distance restraints of 26 Å, and 75% of the identified PPIs reported by the STRING database were with high confidence (scores ≥0.9), confirming the high validity of the identified cross-links for protein structural mapping and PPI analysis. This study demonstrates that O-MICROFASP is a universal and efficient method for proteome-wide XL-MS analysis of microscale samples with high sensitivity and reliability.

交联质谱(XL-MS)是在全球范围内阐明蛋白质结构和蛋白质-蛋白质相互作用(PPIs)的有力工具。然而,由于低丰度交联肽在样品制备过程中会造成严重的样品损失,因此对质量有限的样品进行灵敏的 XL-MS 分析仍然具有挑战性。本文提出了一种优化的微型过滤器辅助样品制备(O-MICROFASP)方法,用于对微量样品进行灵敏的 XL-MS 分析。通过系统地研究和优化关键的实验因素,该方法显著提高了低微克和亚微克样品的 XL 鉴定能力。与传统的 FASP 方法相比,单次分析 1 μg DSS 交联的 HeLa 细胞裂解液(440 对 59)所鉴定出的交联肽多了 7.4 倍。当微反应器中过滤器的表面积进一步减小到 0.058 mm2 时,从 0.5 μg HeLa 细胞裂解液中鉴定出的交联肽的数量增加了 58%。为了加深 XL 蛋白组的鉴定覆盖范围,使用了五种不同类型的交联剂,并用 O-MICROFASP 结合尖端强阳离子交换(SCX)分馏法处理了每微克交联的 HeLa 细胞裂解物。从 5 μg HeLa 细胞裂解液中鉴定出了多达 2741 个独特的交联肽段,代表了 1092 个蛋白质上 2579 个独特的 K-K 连接。约 96% 的蛋白质内交联在 26 Å 的最大距离限制范围内,STRING 数据库报告的 75% 已鉴定的 PPI 具有高置信度(分数≥0.9),证实了已鉴定的交联在蛋白质结构图绘制和 PPI 分析中的高度有效性。这项研究表明,O-MICROFASP 是一种通用、高效的方法,可用于微尺度样品的全蛋白质组 XL-MS 分析,灵敏度高,可靠性强。
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引用次数: 0
Harnessing the Intrinsic Chemical Reactivity of the Mycotoxin Patulin for Immunosensing. 利用霉菌毒素棒曲霉素的内在化学反应性进行免疫传感。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-15 DOI: 10.1021/acs.analchem.4c01631
Hadyn Duncan, Consuelo Agulló, Josep V Mercader, Antonio Abad-Somovilla, Antonio Abad-Fuentes

Mycotoxins are globally pervasive contaminants that threaten food safety worldwide. Regulatory authorities have established maximum permissible levels for certain mycotoxins, and their presence is routinely monitored throughout the food chain to ensure the provision of healthy food and safe feed for humans and animals. While immunoanalytical methods are essential for mycotoxin screening, monoclonal antibodies for the detection of patulin are notably absent. Moreover, leading immunodiagnostic companies currently do not offer rapid tests for patulin in their product portfolios. This deficiency in mycotoxin testing is primarily due to the electrophilic reactivity of patulin. In this study, we exploit this reactivity to develop an innovative strategy that targets the stable adduct formed by the reaction of patulin with aryl-1,2-dithiolates, rather than analyzing the mycotoxin itself. Based on this previously unknown reaction, we present the first collection of monoclonal antibodies, enabling the long-sought goal of sensitive, simple, and user-friendly immunosensing of patulin.

霉菌毒素是全球普遍存在的污染物,威胁着全世界的食品安全。监管机构规定了某些霉菌毒素的最高允许含量,并在整个食物链中对霉菌毒素进行例行监测,以确保为人类和动物提供健康的食品和安全的饲料。虽然免疫分析方法对霉菌毒素筛查至关重要,但用于检测棒曲霉素的单克隆抗体却明显缺乏。此外,主要的免疫诊断公司目前还没有提供针对棒曲霉素的快速检测产品。霉菌毒素检测中的这一不足主要是由于棒曲霉素的亲电反应性造成的。在这项研究中,我们利用这种反应性开发了一种创新策略,以棒曲霉素与芳基-1,2-二硫酸盐反应形成的稳定加合物为目标,而不是分析霉菌毒素本身。基于这种之前未知的反应,我们首次推出了一系列单克隆抗体,从而实现了长期以来一直追求的目标,即对棒曲霉素进行灵敏、简单、易用的免疫传感。
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引用次数: 0
Construction of Near-Infrared Probes with Remarkable Large Stokes Shift Based on a Novel Purine Platform for the Visualization of mtG4 Upregulation during Mitochondrial Disorder in Somatic Cells and Human Sperms. 基于新型嘌呤平台构建具有显著大斯托克斯偏移的近红外探针,用于观察体细胞和人类精子线粒体紊乱过程中的 mtG4 上调。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-15 DOI: 10.1021/acs.analchem.4c01638
Kang-Kang Yu, Kun Li, Hao-Yuan Wang, Xiao-Liang Li, Si-Xian Wu, Wen-Ming Xu, Yan-Hong Liu, Chuan-Fang Wu, Xiao-Qi Yu, Jin-Ku Bao

G-quadruplex structures within the nuclear genome (nG4) is an important regulatory factor, while the function of G4 in the mitochondrial genome (mtG4) still needs to be explored, especially in human sperms. To gain a better understanding of the relationship between mtG4 and mitochondrial function, it is crucial to develop excellent probes that can selectively visualize and track mtG4 in both somatic cells and sperms. Herein, based on our previous research on purine frameworks, we attempted for the first time to extend the conjugated structure from the C-8 site of purine skeleton and discovered that the purine derivative modified by the C-8 aldehyde group is an ideal platform for constructing near-infrared probes with extremely large Stokes shift (>220 nm). Compared with the compound substituted with methylpyridine (PAP), the molecule substituted with methylthiazole orange (PATO) showed better G4 recognition ability, including longer emission (∼720 nm), more significant fluorescent enhancement (∼67-fold), lower background, and excellent photostability. PATO exhibited a sensitive response to mtG4 variation in both somatic cells and human sperms. Most importantly, PATO helped us to discover that mtG4 was significantly increased in cells with mitochondrial respiratory chain damage caused by complex I inhibitors (6-OHDA and rotenone), as well as in human sperms that suffer from oxidative stress. Altogether, our study not only provides a novel ideal molecular platform for constructing high-performance probes but also develops an effective tool for studying the relationship between mtG4 and mitochondrial function in both somatic cells and human sperms.

核基因组(nG4)中的G-四重结构是一个重要的调控因子,而线粒体基因组(mtG4)中G4的功能仍有待探索,尤其是在人类精子中。为了更好地了解 mtG4 与线粒体功能之间的关系,开发出能在体细胞和精子中选择性地观察和追踪 mtG4 的优秀探针至关重要。在此,我们基于之前对嘌呤框架的研究,首次尝试从嘌呤骨架的 C-8 位点扩展共轭结构,发现经 C-8 醛基修饰的嘌呤衍生物是构建具有超大斯托克斯位移(>220 nm)的近红外探针的理想平台。与甲基吡啶(PAP)取代的化合物相比,甲基噻唑橙(PATO)取代的分子具有更好的 G4 识别能力,包括更长的发射时间(∼720 nm)、更显著的荧光增强(∼67 倍)、更低的本底和出色的光稳定性。PATO 对体细胞和人类精子中的 mtG4 变异都有灵敏的反应。最重要的是,PATO 帮助我们发现,在复合体 I 抑制剂(6-OHDA 和鱼藤酮)导致线粒体呼吸链损伤的细胞中,以及在遭受氧化应激的人类精子中,mtG4 显著增加。总之,我们的研究不仅为构建高性能探针提供了一个新的理想分子平台,还为研究体细胞和人类精子中 mtG4 与线粒体功能之间的关系开发了一种有效工具。
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引用次数: 0
High-Throughput Screening for Oligonucleotide Detection by ADE-OPI-MS. 利用 ADE-OPI-MS 进行寡核苷酸检测的高通量筛选。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-14 DOI: 10.1021/acs.analchem.4c02110
Lingling Hu, Zhidan Zhang, Congyu Li, Minghao Han, Mengyao Hao, Xu Zhang, Nida Ahmed, Jianmei Luo, Xiaoyun Lu, Jibin Sun, Huifeng Jiang

Oligonucleotides represent a class of shorter DNA or RNA nucleic acid polymers extensively applied in the biomedical field. Despite progress in detecting and analyzing oligonucleotides, high-throughput analysis of the samples remains challenging. In this work, a high-throughput analysis method for oligonucleotide analysis was developed based on acoustic droplet ejection-open port interface-mass spectrometry (ADE-OPI-MS) technology. This approach was applied to determine the enzymatic activity of terminal deoxynucleotide transferase (TdT) for DNA synthesis, with a rate of 3 s/sample, which enhanced single-sample analysis efficiency approximately 60-fold over the previous gel analysis. After testing approximately 10,000 TdT mutants, we obtained three new variants with higher catalytic activities. Finally, by integrating these mutants, the catalytic activity of TdT was improved about 4 times compared to the starting mutant. Our results successfully established a high-throughput screening method for oligonucleotide analysis, which not only provides a foundation to engineer highly efficient TdT for ab initio synthesis of DNA but also paves the way for the potential application of oligonucleotide analysis in biomedical fields.

寡核苷酸是一类较短的 DNA 或 RNA 核酸聚合物,广泛应用于生物医学领域。尽管在检测和分析寡核苷酸方面取得了进展,但对样本进行高通量分析仍具有挑战性。在这项工作中,基于声学液滴喷射-开放端口接口-质谱(ADE-OPI-MS)技术,开发了一种用于寡核苷酸分析的高通量分析方法。该方法被用于测定 DNA 合成过程中末端脱氧核苷酸转移酶(TdT)的酶活性,分析速度为 3 秒/样品,与之前的凝胶分析相比,单样品分析效率提高了约 60 倍。在测试了约 10,000 个 TdT 突变体后,我们获得了三个催化活性更高的新变体。最后,通过整合这些突变体,TdT 的催化活性比初始突变体提高了约 4 倍。我们的研究结果成功地建立了寡核苷酸分析的高通量筛选方法,不仅为设计高效的 TdT 用于 DNA 的初始合成奠定了基础,而且为寡核苷酸分析在生物医学领域的潜在应用铺平了道路。
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引用次数: 0
Selenium Supplementation Sensor Based on Direct Electrochemistry of Urinary Selenosugar and Total Selenium. 基于尿硒糖和总硒直接电化学的硒补充传感器
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-14 DOI: 10.1021/acs.analchem.4c02391
Meiyan Song, Jialiang Chen, Jingyi Si, Tiantian Man, Qunyan Yao, Fulin Zhu, Fujin Lv, Yuhao Piao, Ying Wan, Changfeng Zhu, Shengyuan Deng

Emerging point-of-care testing methods are extremely beneficial for personalized assessments of trace element metabolism including selenium (Se). Given the lack of timely evaluation methods for well-received Se fortification, an electrochemical solution was developed based on the recently identified urinary selenosugar (Sel) as a marker. The Se content of crude urine was rapidly determined (∼5 min), and the square-wave voltammetric responses of a Se-selective probe (SeSE) composed of liquid metal amalgam demonstrated comparable performance (e.g., detection limit: 19 nM) to central lab benchtop equipment within the physiological range. Meanwhile, SeSE enabled total urinary Se detection via a mere one-step oxidation. Additionally, SeSE was utilized to jointly assess the apparent internalization and utilization rate of two typical nutrients, selenite and selenomethionine, in a rat nutrition model, demonstrating consistent results with those obtained by HPLC-MS and ICP-MS. Upon systematic standardization directed by Ramaley's theory, SeSE was integrated into a battery-operated portable kit (dubbed "SeEye") with a micro electrochemical drive and tablet PC console for one-stop service trials in a local commercial scenario. This study establishes (1) a nutritive value classifier in a low-cost consumer electronic format and (2) noninvasive diagnostic technology for Se supplementation.

新兴的护理点检测方法对于包括硒(Se)在内的微量元素代谢的个性化评估极为有益。鉴于缺乏对广为接受的硒强化剂进行及时评估的方法,我们开发了一种基于最近确定的尿硒糖(Sel)作为标记物的电化学解决方案。粗尿中的硒含量可快速测定(5 分钟),由液态金属汞合金组成的硒选择性探针(SeSE)的方波伏安响应在生理范围内表现出与中央实验室台式设备相当的性能(如检测限:19 nM)。同时,SeSE 只需一步氧化就能实现尿液中总 Se 的检测。此外,SeSE 还被用于在大鼠营养模型中联合评估亚硒酸盐和硒蛋氨酸这两种典型营养物质的表观内化和利用率,结果与 HPLC-MS 和 ICP-MS 获得的结果一致。在拉马利理论指导下进行系统标准化后,SeSE 被集成到一个电池操作的便携式工具包(称为 "SeEye")中,该工具包带有微型电化学驱动器和平板电脑控制台,可在本地商业场景中进行一站式服务试验。这项研究建立了(1)低成本消费电子格式的营养价值分类器和(2)Se 补充的无创诊断技术。
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引用次数: 0
SMMP: A Deep-Coverage Marine Metaproteome Method for Microbial Community Analysis throughout the Water Column Using 1 L of Seawater. SMMP:使用 1 升海水对整个水柱进行微生物群落分析的深度覆盖海洋元蛋白组方法。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-13 DOI: 10.1021/acs.analchem.4c02079
Songduo Wang, Zenghu Zhang, Kaiguang Yang, Jiulong Zhao, Weijie Zhang, Zhiting Wang, Zhen Liang, Yongyu Zhang, Yukui Zhang, Jianhui Liu, Lihua Zhang

Marine microbes drive pivotal transformations in planetary-scale elemental cycles and have crucial impacts on global biogeochemical processes. Metaproteomics is a powerful tool for assessing the metabolic diversity and function of marine microbes. However, hundreds of liters of seawater are required for normal metaproteomic analysis due to the sparsity of microbial populations in seawater, which poses a substantial challenge to the widespread application of marine metaproteomics, particularly for deep seawater. Herein, a sensitive marine metaproteomics workflow, named sensitive marine metaproteome analysis (SMMP), was developed by integrating polycarbonate filter-assisted microbial enrichment, solid-phase alkylation-based anti-interference sample preparation, and narrow-bore nanoLC column for trace peptide separation and characterization. The method provided more than 8500 proteins from 1 L of bathypelagic seawater samples, which covered diverse microorganisms and crucial functions, e.g., the detection of key enzymes associated with the Wood-Ljungdahl pathway. Then, we applied SMMP to investigate vertical variations in the metabolic expression patterns of marine microorganisms from the euphotic zone to the bathypelagic zone. Methane oxidation and carbon monoxide (CO) oxidation were active processes, especially in the bathypelagic zone, which provided a remarkable energy supply for the growth and proliferation of heterotrophic microorganisms. In addition, marker protein profiles detected related to ammonia transport, ammonia oxidation, and carbon fixation highlighted that Thaumarchaeota played a critical role in primary production based on the coupled carbon-nitrogen process, contributing to the storage of carbon and nitrogen in the bathypelagic regions. SMMP has low microbial input requirements and yields in-depth metaproteome analysis, making it a prospective approach for comprehensive marine metaproteomic investigations.

海洋微生物驱动着地球尺度元素循环中的关键转化,对全球生物地球化学过程有着至关重要的影响。元蛋白质组学是评估海洋微生物代谢多样性和功能的有力工具。然而,由于海水中微生物种群稀少,正常的元蛋白质组分析需要数百升海水,这给海洋元蛋白质组学的广泛应用,尤其是深层海水的应用带来了巨大挑战。在此研究中,通过整合聚碳酸酯过滤器辅助微生物富集、基于固相烷基化的抗干扰样品制备以及用于痕量肽分离和表征的窄孔纳米液相色谱柱,开发了一种灵敏的海洋元蛋白质组学工作流程,命名为灵敏的海洋元蛋白质组分析(SMMP)。该方法从 1 升深海海水样品中获得了 8500 多种蛋白质,涵盖了多种微生物和关键功能,如检测与伍德-荣格达尔途径相关的关键酶。然后,我们应用 SMMP 研究了海洋微生物代谢表达模式从透光区到深海区的垂直变化。甲烷氧化和一氧化碳(CO)氧化是活跃的过程,尤其是在深海区,这为异养微生物的生长和增殖提供了显著的能量供应。此外,检测到的与氨转运、氨氧化和碳固定有关的标记蛋白质图谱突出表明,基于碳-氮耦合过程,Thaumarchaeota 在初级生产中发挥了关键作用,有助于在深海区储存碳和氮。SMMP对微生物的投入要求低,并能产生深入的元蛋白组分析,因此是海洋元蛋白组综合研究的一种前瞻性方法。
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引用次数: 0
Modulating Visible-Light Driven NIR Lanthanide Polymer Photocatalysis for Amplification Detection of Exosomal Proteins and Cancer Diagnosis. 调节可见光驱动的近红外镧系聚合物光催化技术,用于外泌体蛋白的放大检测和癌症诊断。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-13 DOI: 10.1021/acs.analchem.4c02168
Haiyan Li, Yafei Chen, Qing Gao, Nan Wang, Ting Yang, Cheng Du, Mingli Chen, Jianhua Wang

Near-infrared (NIR) luminescent lanthanide materials hold great promise for bioanalysis, as they have anti-interference properties. The approach of efficient luminescence is sensitization through a reasonable chromophore to overcome the obstacle of the aqueous phase. The involvement of the surfactant motif is an innovative strategy to arrange the amphiphilic groups to be regularly distributed near the polymer to form a closed sensitized space. Herein, a lanthanide polymer (TCPP-PEI70K-FITC@Yb/SDBS) is designed in which the meso-tetra(4-carboxyphenyl)porphine (TCPP) ligand serves as both a sensitizer and photocatalytic switch. The surfactant sodium dodecyl benzenesulfonate (SDBS) wraps the photosensitive polymers to form a hydrophobic layer, which augments the light-harvesting ability and expedites its photocatalysis. TCPP-PEI70K-FITC@Yb/SDBS is subsequently applied as an amplified photocatalysis toolbox for universally regulating the generation of reactive oxygen species (ROS). Boosting 3,3',5,5'-tetramethylbenzidine (TMB) oxidation to produce blue products, a dual-mode biosensor is fabricated for improving the diagnosis of programmed death ligand-1-positive (PDL1) cancer exosomes. Exosomes were captured by Fe3O4 modified by the PDL1 aptamer, enabling replacement of alkaline phosphatase (ALP)-labeled multiple hybridized chains; then, the isolated ALP triggered a hydrolysis reaction to block the generation of oxTMB. Detection sensitivity improves by 1 order of magnitude through SDBS modulation, down to 104 particles/mL. The sensor performed well clinically in distinguishing cancer patients from healthy individuals, expanding physiological applications of near-infrared lanthanide luminescence.

近红外(NIR)发光镧系元素材料具有抗干扰特性,因此在生物分析领域大有可为。高效发光的方法是通过合理的发色团进行敏化,以克服水相的障碍。表面活性剂图案的参与是一种创新策略,可将两亲基团有规律地分布在聚合物附近,形成一个封闭的敏化空间。本文设计了一种镧系聚合物(TCPP-PEI70K-FITC@Yb/SDBS),其中的中-四(4-羧基苯基)卟吩(TCPP)配体既是敏化剂,又是光催化开关。表面活性剂十二烷基苯磺酸钠(SDBS)将光敏聚合物包裹起来,形成疏水层,从而增强了光收集能力,加快了光催化过程。随后,TCPP-PEI70K-FITC@Yb/SDBS 被用作一种放大的光催化工具箱,用于普遍调节活性氧(ROS)的生成。通过促进 3,3',5,5'-四甲基联苯胺(TMB)氧化产生蓝色产物,制备了一种双模式生物传感器,用于改善程序性死亡配体-1 阳性(PDL1)癌症外泌体的诊断。外泌体被 PDL1 合体修饰的 Fe3O4 捕获,从而取代了碱性磷酸酶(ALP)标记的多重杂交链;然后,分离出的 ALP 触发水解反应,阻止 oxTMB 的生成。通过 SDBS 调制,检测灵敏度提高了 1 个数量级,低至 104 个颗粒/毫升。该传感器在区分癌症患者和健康人方面的临床表现良好,拓展了近红外镧系元素发光的生理应用。
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引用次数: 0
CRISPR/Cas13a-Responsive and RNA-Bridged DNA Hydrogel Capillary Sensor for Point-of-Care Detection of RNA. 用于 RNA 床旁检测的 CRISPR/Cas13a 反应性和 RNA 桥接 DNA 水凝胶毛细管传感器。
IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Pub Date : 2024-07-13 DOI: 10.1021/acs.analchem.4c02087
Hui Wang, Honghong Wang, Hongru Pian, Fengxia Su, Fu Tang, Desheng Chen, Junjie Chen, Yongqiang Wen, X Chris Le, Zhengping Li

Disease diagnostics and surveillance increasingly highlight the importance of portable, cost-effective, and sensitive point-of-care (POC) detection of nucleic acids. Here, we report a CRISPR/Cas13a-responsive and RNA-bridged DNA hydrogel capillary sensor for the direct and visual detection of specific RNA with high sensitivity. The capillary sensor was simply prepared by loading RNA-cross-linking DNA hydrogel film (∼0.2 mm ± 0.02 mm) at the end of a capillary. When CRISPR/Cas13a specifically recognizes the target RNA, the RNA bridge in the hydrogel film is cleaved by the trans-cleavage activity of CRISPR/Cas13a, increasing the permeability of the hydrogel film. Different concentrations of target RNA activate different amounts of Cas13a, cleaving different amounts of the RNA bridge in the hydrogel and causing corresponding changes in the permeability of the hydrogel. Therefore, samples containing different amounts of the target RNA travel to different distances in the capillary. Visual reading of the distance provides quantitative detection of the RNA target without the need for any nucleic acid amplification or auxiliary equipment. The technique was successfully used for the determination of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in clinical nasopharyngeal (NP) swab and saliva samples. Easily quantifiable distance using a ruler eliminates the need for any optical or electrochemical detection equipment, making this assay potentially useful for POC and on-site applications.

疾病诊断和监测日益凸显出便携、经济、灵敏的床旁核酸检测(POC)的重要性。在这里,我们报告了一种 CRISPR/Cas13a 响应和 RNA 桥接 DNA 水凝胶毛细管传感器,可直接可视地高灵敏度检测特定 RNA。毛细管传感器的制备非常简单,只需在毛细管末端加载 RNA 交联 DNA 水凝胶薄膜(∼0.2 mm ± 0.02 mm)。当 CRISPR/Cas13a 特异性识别目标 RNA 时,水凝胶膜中的 RNA 桥会被 CRISPR/Cas13a 的反式裂解活性裂解,从而增加水凝胶膜的渗透性。不同浓度的目标 RNA 会激活不同量的 Cas13a,裂解水凝胶中不同量的 RNA 桥,导致水凝胶的渗透性发生相应的变化。因此,含有不同量目标 RNA 的样品在毛细管中的移动距离也不同。通过目视读取距离可以定量检测目标 RNA,而无需任何核酸扩增或辅助设备。该技术已成功用于测定临床鼻咽(NP)拭子和唾液样本中的严重急性呼吸系统综合征冠状病毒 2(SARS-CoV-2)RNA。使用尺子即可轻松量化距离,无需任何光学或电化学检测设备,因此这种检测方法可用于 POC 和现场应用。
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引用次数: 0
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Analytical Chemistry
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