Cellular and Molecular Biotechnology Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Japan.
Methods Mol Biol. 2024;2763:151-158. doi: 10.1007/978-1-0716-3670-1_13.
The large variety and high concentration of O-glycans are characteristic properties of mucins and play a crucial role in their unique functions. Analyzing the O-glycans of mucins is essential for investigating the functions of mucins. Eliminative oximation is an aqueous reaction that can be used to obtain O-glycan oximes from mucins. Using diazabicyclo undec-7ene (DBU) as a base, an organic superbase that can be removed with an organic solvent during solid-phase extraction, and adding hydroxylamine to the reaction mixture in advance, the O-glycans released from the mucin are immediately converted to the corresponding glycan oximes. The glycan oxime can then be fluorescently labeled with a fluorescent labeling reagent and 2-picoline borane via reductive amination. O-glycans that have been fluorescently labeled can be analyzed using conventional HPLC techniques.
O-聚糖的种类繁多且浓度高是黏蛋白的特征属性，对其独特功能起着至关重要的作用。分析黏蛋白的 O-聚糖对于研究黏蛋白的功能至关重要。消除性肟化是一种可以从黏蛋白中获得 O-聚糖肟的水相反应。使用 1,8-二氮杂二环十一碳-7-烯（DBU）作为碱，它是一种有机超强碱，可在固相萃取过程中用有机溶剂去除，并在反应混合物中预先添加羟胺，从黏蛋白中释放的 O-聚糖可立即转化为相应的聚糖肟。然后，聚糖肟可通过还原胺化与荧光标记试剂和 2-吡啶硼烷反应进行荧光标记。经荧光标记的 O-聚糖可使用常规 HPLC 技术进行分析。
