{"title":"毛细管电泳与荧光各向异性竞争免疫测定法检测胰高血糖素的比较。","authors":"Yao Wang, Emily L. Skinner, Michael G. Roper","doi":"10.1002/elps.202400080","DOIUrl":null,"url":null,"abstract":"<p>Glucagon plays a crucial role in regulating glucose homeostasis; unfortunately, the mechanisms controlling its release are still unclear. Capillary electrophoresis (CE)- and fluorescence anisotropy (FA)-immunoassays (IA) have been used for online measurements of hormone secretion on microfluidic platforms, although their use in glucagon assays is less common. We set out to compare a glucagon-competitive IA using these two techniques. Theoretical calibration curves were generated for both CE- and FA–IA and results indicated that CE-IA provided higher sensitivity than FA–IA. These results were confirmed in an experiment where both assays showed limits of detection (LOD) of 30 nM, but the CE-IA had ∼300-fold larger sensitivity from 0 to 200 nM glucagon. However, in online experiments where reagents were mixed within the device, the sensitivity of the CE-IA was reduced ∼3-fold resulting in a higher LOD of 70 nM, whereas the FA–IA remained essentially unchanged. This lowered sensitivity in the online CE-IA was likely due to poor sampling by electroosmotic flow from the high salt solution necessary in online experiments, whereas pressure-based sampling used in FA–IA was not affected. We conclude that FA–IA, despite lowered sensitivity, is more suitable for online mixing scenarios due to the ability to use pressure-driven flow and other practical advantages such as the use of larger channels.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"45 19-20","pages":"1692-1700"},"PeriodicalIF":3.0000,"publicationDate":"2024-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Comparison between capillary electrophoresis and fluorescence anisotropy competitive immunoassay for glucagon\",\"authors\":\"Yao Wang, Emily L. Skinner, Michael G. Roper\",\"doi\":\"10.1002/elps.202400080\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>Glucagon plays a crucial role in regulating glucose homeostasis; unfortunately, the mechanisms controlling its release are still unclear. Capillary electrophoresis (CE)- and fluorescence anisotropy (FA)-immunoassays (IA) have been used for online measurements of hormone secretion on microfluidic platforms, although their use in glucagon assays is less common. We set out to compare a glucagon-competitive IA using these two techniques. Theoretical calibration curves were generated for both CE- and FA–IA and results indicated that CE-IA provided higher sensitivity than FA–IA. These results were confirmed in an experiment where both assays showed limits of detection (LOD) of 30 nM, but the CE-IA had ∼300-fold larger sensitivity from 0 to 200 nM glucagon. However, in online experiments where reagents were mixed within the device, the sensitivity of the CE-IA was reduced ∼3-fold resulting in a higher LOD of 70 nM, whereas the FA–IA remained essentially unchanged. This lowered sensitivity in the online CE-IA was likely due to poor sampling by electroosmotic flow from the high salt solution necessary in online experiments, whereas pressure-based sampling used in FA–IA was not affected. We conclude that FA–IA, despite lowered sensitivity, is more suitable for online mixing scenarios due to the ability to use pressure-driven flow and other practical advantages such as the use of larger channels.</p>\",\"PeriodicalId\":11596,\"journal\":{\"name\":\"ELECTROPHORESIS\",\"volume\":\"45 19-20\",\"pages\":\"1692-1700\"},\"PeriodicalIF\":3.0000,\"publicationDate\":\"2024-07-10\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"ELECTROPHORESIS\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://onlinelibrary.wiley.com/doi/10.1002/elps.202400080\",\"RegionNum\":3,\"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":"ELECTROPHORESIS","FirstCategoryId":"99","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1002/elps.202400080","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
Comparison between capillary electrophoresis and fluorescence anisotropy competitive immunoassay for glucagon
Glucagon plays a crucial role in regulating glucose homeostasis; unfortunately, the mechanisms controlling its release are still unclear. Capillary electrophoresis (CE)- and fluorescence anisotropy (FA)-immunoassays (IA) have been used for online measurements of hormone secretion on microfluidic platforms, although their use in glucagon assays is less common. We set out to compare a glucagon-competitive IA using these two techniques. Theoretical calibration curves were generated for both CE- and FA–IA and results indicated that CE-IA provided higher sensitivity than FA–IA. These results were confirmed in an experiment where both assays showed limits of detection (LOD) of 30 nM, but the CE-IA had ∼300-fold larger sensitivity from 0 to 200 nM glucagon. However, in online experiments where reagents were mixed within the device, the sensitivity of the CE-IA was reduced ∼3-fold resulting in a higher LOD of 70 nM, whereas the FA–IA remained essentially unchanged. This lowered sensitivity in the online CE-IA was likely due to poor sampling by electroosmotic flow from the high salt solution necessary in online experiments, whereas pressure-based sampling used in FA–IA was not affected. We conclude that FA–IA, despite lowered sensitivity, is more suitable for online mixing scenarios due to the ability to use pressure-driven flow and other practical advantages such as the use of larger channels.
期刊介绍:
ELECTROPHORESIS is an international journal that publishes original manuscripts on all aspects of electrophoresis, and liquid phase separations (e.g., HPLC, micro- and nano-LC, UHPLC, micro- and nano-fluidics, liquid-phase micro-extractions, etc.).
Topics include new or improved analytical and preparative methods, sample preparation, development of theory, and innovative applications of electrophoretic and liquid phase separations methods in the study of nucleic acids, proteins, carbohydrates natural products, pharmaceuticals, food analysis, environmental species and other compounds of importance to the life sciences.
Papers in the areas of microfluidics and proteomics, which are not limited to electrophoresis-based methods, will also be accepted for publication. Contributions focused on hyphenated and omics techniques are also of interest. Proteomics is within the scope, if related to its fundamentals and new technical approaches. Proteomics applications are only considered in particular cases.
Papers describing the application of standard electrophoretic methods will not be considered.
Papers on nanoanalysis intended for publication in ELECTROPHORESIS should focus on one or more of the following topics:
• Nanoscale electrokinetics and phenomena related to electric double layer and/or confinement in nano-sized geometry
• Single cell and subcellular analysis
• Nanosensors and ultrasensitive detection aspects (e.g., involving quantum dots, "nanoelectrodes" or nanospray MS)
• Nanoscale/nanopore DNA sequencing (next generation sequencing)
• Micro- and nanoscale sample preparation
• Nanoparticles and cells analyses by dielectrophoresis
• Separation-based analysis using nanoparticles, nanotubes and nanowires.