High-throughput solid-phase permethylation of glycans prior to mass spectrometry
High-throughput solid-phase permethylation of glycans prior to mass spectrometry
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DOI：
10.1002/rcm.3395
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发表时间：
2008-01-01
影响因子：
2
通讯作者：
Novotny, Milos V.
中科院分区：
文献类型：
--
作者：
Kang, Pilsoo;Mechref, Yehia;Novotny, Milos V.
在进行质谱测定之前，聚糖的全甲基化现已成为糖缀合物分析中历史悠久的方法，这是因为其具有同时分析中性和酸性聚糖的优点以及增强的灵敏度和更容易的串联质谱解释。虽然该方法的不同溶剂萃取版本通常足以用于不同的结构研究，但在针对少量分析材料的定量测定中，它们通常不太令人满意。为了克服这些困难，我们最近引入了固相毛细管全甲基化技术（Kang等人，快速通信质谱仪2005; 19：3421）用于微尺度测定。在这里，我们描述了一个非常有用的高通量扩展的固相方法，利用旋转柱填充氢氧化钠珠。该程序已被彻底优化，以匹配先前使用的毛细管技术的分析性能参数。正如人血清的高精度糖组学分析所证明的那样，这种方法学改进提供了简单性和再现性，允许在不到20分钟的时间内完成12-18个样品的完全全甲基化。
Permethylation of glycans prior to their mass spectrometric determination has now become a time-honored methodology in glycoconjugate analysis due to the advantage of a simultaneous analysis of neutral and acidic glycans as well as enhanced sensitivity and easier tandem mass spectrometry interpretation. While the different solvent extraction-based versions of this method often suffice in different structural studies, they are generally less satisfactory in the quantitative determinations aiming at minor quantities of the analyzed materials. To overcome these difficulties, we recently introduced a solid-phase capillary permethylation technique (Kang et al., Rapid Commun. Mass Spectrom. 2005; 19: 3421) for microscale determination. Here, we describe a very useful high-throughput extension of the solid-phase methodology utilizing spin columns packed with sodium hydroxide beads. This procedure has been thoroughly optimized to match the analytical performance parameters of the previously used capillary technique. As demonstrated with a high-precision glycomic profiling analysis of human blood serum, this methodological improvement offers simplicity and reproducibility, allowing the complete permethylation of 12-18 samples in less than 20 min. Copyright (C) 2008 John Wiley & Sons, Ltd.