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。

IF 3 3区 生物学 Q2 BIOCHEMICAL RESEARCH METHODS
Yao Wang, Emily L. Skinner, Michael G. Roper
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引用次数: 0

摘要

胰高血糖素在调节葡萄糖稳态中起着至关重要的作用;遗憾的是,控制其释放的机制仍不清楚。毛细管电泳(CE)和荧光各向异性(FA)免疫测定(IA)已被用于微流控平台上激素分泌的在线测量,但它们在胰高血糖素测定中的应用还不太常见。我们利用这两种技术对胰高血糖素竞争性免疫测定进行了比较。我们生成了 CE-IA 和 FA-IA 的理论校准曲线,结果表明 CE-IA 的灵敏度高于 FA-IA。这些结果在一项实验中得到了证实,在该实验中,两种检测方法的检测限(LOD)均为 30 nM,但 CE-IA 在 0 至 200 nM 胰高血糖素范围内的灵敏度比 FA-IA 高 300 倍。然而,在装置内混合试剂的在线实验中,CE-IA 的灵敏度降低了 3 倍,导致检测限提高到 70 nM,而 FA-IA 则基本保持不变。在线 CE-IA 灵敏度降低的原因可能是在线实验中所需的高盐溶液的电渗流取样效果不佳,而 FA-IA 中使用的基于压力的取样不受影响。我们的结论是,FA-IA 尽管灵敏度较低,但由于能够使用压力驱动的流动和其他实际优势(如使用较大的通道),因此更适合在线混合方案。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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.

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来源期刊
ELECTROPHORESIS
ELECTROPHORESIS 生物-分析化学
CiteScore
6.30
自引率
13.80%
发文量
244
审稿时长
1.9 months
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.
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