Flow-LAMP：使用基于散射的流式细胞术对涡流产生的多分散凝胶珠进行无标签数字LAMP。

IF 6.7 1区化学 Q1 CHEMISTRY, ANALYTICAL

Analytical Chemistry Pub Date : 2025-10-12 DOI:10.1021/acs.analchem.5c04768

Yuchong Zheng,Wanjun Yao,Zerui Wu,Liqun He,Weidong Zheng,Zida Li

{"title":"Flow-LAMP：使用基于散射的流式细胞术对涡流产生的多分散凝胶珠进行无标签数字LAMP。","authors":"Yuchong Zheng,Wanjun Yao,Zerui Wu,Liqun He,Weidong Zheng,Zida Li","doi":"10.1021/acs.analchem.5c04768","DOIUrl":null,"url":null,"abstract":"Accurate nucleic acid quantification is vital for clinical diagnostics, yet the widespread adoption of digital PCR remains limited due to its reliance on fluorescence detection and specialized microfluidics. We present Flow-LAMP, a label-free digital assay integrating loop-mediated isothermal amplification with scatter-based flow cytometric analysis of agarose gel beads. Polydisperse gel beads are formed by vortex emulsification and retain magnesium pyrophosphate precipitate in positive reactions. Flow cytometry enables volume and amplification readouts via forward (FSC) and side scattering (SSC) signals, respectively. We confirmed that SSC was strongly correlated with amplification products, while FSC-Height accurately reflected the bead volume. Using Epstein-Barr virus plasmid, Flow-LAMP achieved accurate quantification with a limit of detection of 38.15 copies/μL. Results from testing clinical plasma samples correlated well with qPCR and digital PCR. By eliminating fluorescent labeling and microfluidics, Flow-LAMP offers a cost-effective and accessible platform for digital nucleic acid detection using standard lab equipment.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"40 1","pages":""},"PeriodicalIF":6.7000,"publicationDate":"2025-10-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Flow-LAMP: Label-Free Digital LAMP Using Scatter-Based Flow Cytometry on Vortex-Generated Polydisperse Gel Beads.\",\"authors\":\"Yuchong Zheng,Wanjun Yao,Zerui Wu,Liqun He,Weidong Zheng,Zida Li\",\"doi\":\"10.1021/acs.analchem.5c04768\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Accurate nucleic acid quantification is vital for clinical diagnostics, yet the widespread adoption of digital PCR remains limited due to its reliance on fluorescence detection and specialized microfluidics. We present Flow-LAMP, a label-free digital assay integrating loop-mediated isothermal amplification with scatter-based flow cytometric analysis of agarose gel beads. Polydisperse gel beads are formed by vortex emulsification and retain magnesium pyrophosphate precipitate in positive reactions. Flow cytometry enables volume and amplification readouts via forward (FSC) and side scattering (SSC) signals, respectively. We confirmed that SSC was strongly correlated with amplification products, while FSC-Height accurately reflected the bead volume. Using Epstein-Barr virus plasmid, Flow-LAMP achieved accurate quantification with a limit of detection of 38.15 copies/μL. Results from testing clinical plasma samples correlated well with qPCR and digital PCR. By eliminating fluorescent labeling and microfluidics, Flow-LAMP offers a cost-effective and accessible platform for digital nucleic acid detection using standard lab equipment.\",\"PeriodicalId\":27,\"journal\":{\"name\":\"Analytical Chemistry\",\"volume\":\"40 1\",\"pages\":\"\"},\"PeriodicalIF\":6.7000,\"publicationDate\":\"2025-10-12\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Analytical Chemistry\",\"FirstCategoryId\":\"92\",\"ListUrlMain\":\"https://doi.org/10.1021/acs.analchem.5c04768\",\"RegionNum\":1,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"CHEMISTRY, ANALYTICAL\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Analytical Chemistry","FirstCategoryId":"92","ListUrlMain":"https://doi.org/10.1021/acs.analchem.5c04768","RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"CHEMISTRY, ANALYTICAL","Score":null,"Total":0}

引用次数: 0

摘要

准确的核酸定量对临床诊断至关重要，但由于其依赖于荧光检测和专门的微流体，数字PCR的广泛采用仍然受到限制。我们提出flow - lamp，这是一种无标记的数字检测方法，将环介导的等温扩增与基于散射的琼脂糖凝胶珠流式细胞分析相结合。通过涡流乳化形成多分散凝胶珠，并在正反应中保留焦磷酸镁沉淀。流式细胞术分别通过正向（FSC）和侧散射（SSC）信号实现体积和放大读数。我们证实，SSC与扩增产物密切相关，而FSC-Height准确反映了珠粒体积。Flow-LAMP采用Epstein-Barr病毒质粒进行定量，检测限为38.15 copies/μL。临床血浆样本检测结果与qPCR和数字PCR具有良好的相关性。通过消除荧光标记和微流体，Flow-LAMP为使用标准实验室设备的数字核酸检测提供了一个具有成本效益和可访问的平台。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

查看原文本刊更多论文

Flow-LAMP: Label-Free Digital LAMP Using Scatter-Based Flow Cytometry on Vortex-Generated Polydisperse Gel Beads.

Accurate nucleic acid quantification is vital for clinical diagnostics, yet the widespread adoption of digital PCR remains limited due to its reliance on fluorescence detection and specialized microfluidics. We present Flow-LAMP, a label-free digital assay integrating loop-mediated isothermal amplification with scatter-based flow cytometric analysis of agarose gel beads. Polydisperse gel beads are formed by vortex emulsification and retain magnesium pyrophosphate precipitate in positive reactions. Flow cytometry enables volume and amplification readouts via forward (FSC) and side scattering (SSC) signals, respectively. We confirmed that SSC was strongly correlated with amplification products, while FSC-Height accurately reflected the bead volume. Using Epstein-Barr virus plasmid, Flow-LAMP achieved accurate quantification with a limit of detection of 38.15 copies/μL. Results from testing clinical plasma samples correlated well with qPCR and digital PCR. By eliminating fluorescent labeling and microfluidics, Flow-LAMP offers a cost-effective and accessible platform for digital nucleic acid detection using standard lab equipment.

求助全文

通过发布文献求助，成功后即可免费获取论文全文。去求助

来源期刊

Analytical Chemistry 化学-分析化学

CiteScore

12.10

自引率

12.20%

发文量

1949

审稿时长

1.4 months

期刊介绍： Analytical Chemistry, a peer-reviewed research journal, focuses on disseminating new and original knowledge across all branches of analytical chemistry. Fundamental articles may explore general principles of chemical measurement science and need not directly address existing or potential analytical methodology. They can be entirely theoretical or report experimental results. Contributions may cover various phases of analytical operations, including sampling, bioanalysis, electrochemistry, mass spectrometry, microscale and nanoscale systems, environmental analysis, separations, spectroscopy, chemical reactions and selectivity, instrumentation, imaging, surface analysis, and data processing. Papers discussing known analytical methods should present a significant, original application of the method, a notable improvement, or results on an important analyte.