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首页> 外文学位 >Off-line sample preparation techniques for improved analysis of biological oligosaccharides by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS).
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Off-line sample preparation techniques for improved analysis of biological oligosaccharides by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS).

机译：离线样品制备技术，用于通过基质辅助激光解吸/电离质谱（MALDI MS）改进对生物寡糖的分析。

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摘要

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) is a powerful analytical tool available for studying different biomolecules and their complex mixtures. Special consideration for sample handling, preparation techniques and subsequent analysis by MALDI MS must be taken in order to guarantee the best analysis possible in terms of accuracy and reproducibility of analyte identification, relative and absolute quantification, and the number of mixture constituents detected and characterized. The use of various compounds as matrix additives was found to enhance ion signal intensities of the analyte biomolecules, in particular carbohydrates, over the conventionally used matrix materials. Aniline and N,N-dimethylaniline were identified as effective additives to 2,5-dihydroxybenzoic acid, a widely used MALDI matrix, for superior analysis of carbohydrates. Perhaps the most important advantages of using these materials is the fact that sample oligosaccharides need not be derivatized, thus making highly sensitive analysis of native glycans attainable by MALDI MS. Use of these materials also offers some unique advantages, such as a more reliable and reproducible quantitative analysis of sugars and a possibility for designing efficient workflows for automated detection of oligosaccharides in complex biological samples. Multidimensional separation procedures, involving various types of liquid chromatography and capillary electrophoresis, were employed for targeted sample fractionation according to modifications of interest, as well as to reduce the signal suppression of the less abundant analytes and to improve the resolution of the sample mixture components. Capillary electrophoresis was demonstrated to be an effective technique for isolating glycopeptides containing sialic acids. Cellulose-based solid phase extraction procedures were found to be extremely useful and efficient for isolation of glycopeptides, and were shown to offer real practical benefits for routine glycoproteomic and glycomic applications. Reversed-phase chromatography was proven to be essential in oligosaccharide purification and in site-specific analysis of protein glycosylation. Examples of the aforementioned techniques and methods with respect to their application to various controlled standard systems, as well as real-life precious biological samples are presented in this work.
机译：基质辅助激光解吸/电离质谱（MALDI MS）是一种功能强大的分析工具，可用于研究不同的生物分子及其复杂混合物。必须特别考虑样品处理，制备技术和MALDI MS进行的后续分析，以确保在分析物鉴定的准确性和可重复性，相对和绝对定量以及检测和表征的混合物成分的数量方面进行最佳分析。发现使用各种化合物作为基质添加剂比常规使用的基质材料增强了分析物生物分子，特别是碳水化合物的离子信号强度。苯胺和N，N-二甲基苯胺被认为是2,5-二羟基苯甲酸（一种广泛使用的MALDI基质）的有效添加剂，可对碳水化合物进行出色的分析。使用这些材料的最重要的优势也许是不需要将样品中的寡糖衍生化的事实，因此可以通过MALDI MS实现对天然聚糖的高灵敏度分析。这些材料的使用还提供了一些独特的优势，例如对糖进行更可靠和可重现的定量分析，并有可能设计出有效的工作流程来自动检测复杂生物样品中的寡糖。多维分离程序涉及各种类型的液相色谱和毛细管电泳，可根据感兴趣的修改用于目标样品的分馏，并减少对含量较低的分析物的信号抑制，并提高样品混合物组分的分辨率。毛细管电泳被证明是一种分离含有唾液酸的糖肽的有效技术。发现基于纤维素的固相萃取程序对于糖肽的分离非常有用和高效，并且显示出对常规糖蛋白组学和糖组学应用具有真正的实际益处。事实证明，反相色谱法对寡糖纯化和蛋白质糖基化的位点特异性分析至关重要。在这项工作中介绍了上述技术和方法在应用于各种受控标准系统以及现实生活中宝贵的生物样品方面的示例。

著录项

  • 作者

    Snovida, Sergei I.;

  • 作者单位

    University of Manitoba (Canada).;

  • 授予单位 University of Manitoba (Canada).;
  • 学科 Biology Molecular.;Chemistry Biochemistry.;Chemistry Analytical.
  • 学位 Ph.D.
  • 年度 2010
  • 页码 172 p.
  • 总页数 172
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

  • 入库时间 2022-08-17 11:36:56

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