Optimized Workflow for In-depth N-glycosylation Analysis of 2D and 3D Breast Cancer Cell Cultures and Xenografted Tumors

Native intact N-glycopeptides analysis can access to a lot of useful information including N-glycosites, N-glycans and N-glycoproteins of original complex samples. The traditional sample preprocessing method is to analyze the tryptic intact N-glycopeptides enriched using hydrophilic materials or some new materials by LC-MS/MS. However, the number of identified intact N-glycopeptides was very limited using existing methods. Herein, we systematically compared the different analytical methods, including the use of different sources of trypsin, combinations of different proteases and different enrichment materials.