Audrey P. Luu , Shreedevi S. Rao , Humza Y. Malik, Robin B. Shi, Adam A. Toubian, Daniel T. Kamei
{"title":"研究利用侧流免疫测定从两相水溶液系统中进行底相萃取以检测细菌。","authors":"Audrey P. Luu , Shreedevi S. Rao , Humza Y. Malik, Robin B. Shi, Adam A. Toubian, Daniel T. Kamei","doi":"10.1016/j.ab.2024.115634","DOIUrl":null,"url":null,"abstract":"<div><p>Lateral-flow immunoassays (LFAs) can be used to diagnose urinary tract infections caused by <em>Escherichia coli</em> (<em>E. coli</em>) at the point of care. Unfortunately, urine samples containing dilute concentrations of <em>E. coli</em> can yield false negative results on LFAs. Our laboratory was first to implement aqueous two-phase systems (ATPSs) to preconcentrate samples into smaller volumes prior to their application on LFAs. This is achieved by manipulating the ratio of the volume of the top phase to that of the bottom phase (volume ratio; VR) and concentrating biomarkers in the bottom phase which, when applied to LFAs in fixed volumes, leads to corresponding improvements in sensitivity. This work is the first demonstration that the same LOD can be achieved irrespective of the VR when the entire bottom phase is added to LFAs. A custom 3D-printed device was also developed to decrease liquid handling steps. Across different VRs expected from patient urine variability, this diagnostic workflow successfully detected <em>E. coli</em> concentrations down to 2 × 10<sup>5</sup> colony-forming units (cfu) mL<sup>−1</sup> in synthetic urine, demonstrating consistent 10-fold improvements in sensitivity compared to trials conducted without ATPS preconcentration. This method successfully addresses the variability of patient samples while remaining easy to use at the point of care.</p></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"694 ","pages":"Article 115634"},"PeriodicalIF":2.6000,"publicationDate":"2024-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Investigating bottom phase extraction from aqueous two-phase systems for detecting bacteria using the lateral-flow immunoassay\",\"authors\":\"Audrey P. Luu , Shreedevi S. Rao , Humza Y. Malik, Robin B. Shi, Adam A. Toubian, Daniel T. Kamei\",\"doi\":\"10.1016/j.ab.2024.115634\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Lateral-flow immunoassays (LFAs) can be used to diagnose urinary tract infections caused by <em>Escherichia coli</em> (<em>E. coli</em>) at the point of care. Unfortunately, urine samples containing dilute concentrations of <em>E. coli</em> can yield false negative results on LFAs. Our laboratory was first to implement aqueous two-phase systems (ATPSs) to preconcentrate samples into smaller volumes prior to their application on LFAs. This is achieved by manipulating the ratio of the volume of the top phase to that of the bottom phase (volume ratio; VR) and concentrating biomarkers in the bottom phase which, when applied to LFAs in fixed volumes, leads to corresponding improvements in sensitivity. This work is the first demonstration that the same LOD can be achieved irrespective of the VR when the entire bottom phase is added to LFAs. A custom 3D-printed device was also developed to decrease liquid handling steps. Across different VRs expected from patient urine variability, this diagnostic workflow successfully detected <em>E. coli</em> concentrations down to 2 × 10<sup>5</sup> colony-forming units (cfu) mL<sup>−1</sup> in synthetic urine, demonstrating consistent 10-fold improvements in sensitivity compared to trials conducted without ATPS preconcentration. This method successfully addresses the variability of patient samples while remaining easy to use at the point of care.</p></div>\",\"PeriodicalId\":7830,\"journal\":{\"name\":\"Analytical biochemistry\",\"volume\":\"694 \",\"pages\":\"Article 115634\"},\"PeriodicalIF\":2.6000,\"publicationDate\":\"2024-07-31\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Analytical biochemistry\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0003269724001787\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Analytical biochemistry","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0003269724001787","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
Investigating bottom phase extraction from aqueous two-phase systems for detecting bacteria using the lateral-flow immunoassay
Lateral-flow immunoassays (LFAs) can be used to diagnose urinary tract infections caused by Escherichia coli (E. coli) at the point of care. Unfortunately, urine samples containing dilute concentrations of E. coli can yield false negative results on LFAs. Our laboratory was first to implement aqueous two-phase systems (ATPSs) to preconcentrate samples into smaller volumes prior to their application on LFAs. This is achieved by manipulating the ratio of the volume of the top phase to that of the bottom phase (volume ratio; VR) and concentrating biomarkers in the bottom phase which, when applied to LFAs in fixed volumes, leads to corresponding improvements in sensitivity. This work is the first demonstration that the same LOD can be achieved irrespective of the VR when the entire bottom phase is added to LFAs. A custom 3D-printed device was also developed to decrease liquid handling steps. Across different VRs expected from patient urine variability, this diagnostic workflow successfully detected E. coli concentrations down to 2 × 105 colony-forming units (cfu) mL−1 in synthetic urine, demonstrating consistent 10-fold improvements in sensitivity compared to trials conducted without ATPS preconcentration. This method successfully addresses the variability of patient samples while remaining easy to use at the point of care.
期刊介绍:
The journal''s title Analytical Biochemistry: Methods in the Biological Sciences declares its broad scope: methods for the basic biological sciences that include biochemistry, molecular genetics, cell biology, proteomics, immunology, bioinformatics and wherever the frontiers of research take the field.
The emphasis is on methods from the strictly analytical to the more preparative that would include novel approaches to protein purification as well as improvements in cell and organ culture. The actual techniques are equally inclusive ranging from aptamers to zymology.
The journal has been particularly active in:
-Analytical techniques for biological molecules-
Aptamer selection and utilization-
Biosensors-
Chromatography-
Cloning, sequencing and mutagenesis-
Electrochemical methods-
Electrophoresis-
Enzyme characterization methods-
Immunological approaches-
Mass spectrometry of proteins and nucleic acids-
Metabolomics-
Nano level techniques-
Optical spectroscopy in all its forms.
The journal is reluctant to include most drug and strictly clinical studies as there are more suitable publication platforms for these types of papers.