Monoclonal antibodies( mAbs ) are efficacious therapeutic agents against various complex diseases.N-glycosylation characterization of mAbs is based primarily on fluorescence derivatization coupled with hydrophilic interaction liquid chromatography( HILIC ).However, this method has some drawbacks.Presently,the N-glycosylation of mAbs was analyzed by methylamidation and silica-hydride-based normal phase chromatography( SiH-NPC ).Samples were analyzed by liquid chromatography-mass spectrometry after enzymatic cleavage, methylamidation, and purification.SiH-NPC had some advantages compared to HILIC.It offered a novel separation mechanism that allowed for high resolution using salt-free mobile phases,avoiding contamination of the mass spectrometer.SiH-NPC may be applicable for rapid analysis.Its structure was more stable than that of HILIC,and it provided a long service life.In combination with the sialic acid derivatization,SiH-NPC presented a significant advantage in the analysis of sialylated glycans as well as isomeric oligosaccharides by liquid chromatography-mass spectrometry.The approach has potential applications in the biopharmaceutical industry.%采用甲胺化衍生结合基于硅氢化物固定相的正相色谱( SiH-NPC)分析单抗的 N-糖基化.样品经酶切、甲胺化衍生、纯化后由液相色谱-质谱进行分析.结果表明,相较于亲水相互作用色谱( HILIC),SiH-NPC 分离机制不同,使用常规的无盐流动相即可实现高分离度,避免污染质谱,色谱柱结构稳定,使用寿命长,更适合快速分析.结合唾液酸衍生方法,SiH-NPC在液相色谱-质谱联用鉴定酸性糖和糖异构体方面呈现显著优势,在生物制药行业中具有重要的应用潜力.
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