Updated project metadata. N-linked glycoproteins are known to be important for spermatozoa development and gametes fusion. However, little is known about detailed information of these glycoproteins, particularly the glycosylation sites and its corresponding glycan structures in human spermatozoa. In present study, an in house software coupled with LC-MS/MS analysis of intact glycopeptides was employed to establish a large scale of site-specific N-glycoproteome map in human spermatozoa. The results mapped the large-scale N-glycoproteins in human spermatozoa and interpreted the precise glycan structures with the corresponding glycosites. These presented data laid the foundation for the exploration of biological function of glycosylation on human spermatozoa and glycan structure involved male infertility.