Protein glycosylation plays essential roles in regulating innate and adaptive immune response. Previous studies only focused on individual protein-glycan interactions or specific glycoform changes during T cell activation, yet the systematic characterization of protein glycosylation alterations remains insufficiently elucidated. To address these limitations, we conducted temporally resolved quantitative analysis of glycoforms, site-specific glycans, glycoproteins, and glycosylation enzymes in activated Jurkat T cells, and successfully portrayed the dynamic landscape of protein glycosylation during Jurkat T cell activation. We found the heterogeneity and number of significantly upregulated glycopeptides increased along with activation. For most glycopeptides, their alteration patterns did not correlate with the abundance of their glycoprotein substrates. However, functional molecules including CD69, CD28, and PTPRC demonstrated co-upregulation at both the protein and glycosylation levels. Correlation analysis between glycopeptides and glycotransferases indicated that sialylated or fucosylated peptides were well correlated with enzymes involved in glycan branching and capping. Comparative analysis of global peptides, glycopeptides, and phosphopeptides revealed their distinctive changing patterns along Jurkat T cell activation, and only glycosylation demonstrated a steady increase trend with a large proportion of upregulated glycopeptides. Collectively, this integrated multi-proteomics characterization of activated Jurkat T cells provided insights for the development of novel therapeutic strategy targeting glycosylation.