Abstract The Chinese pangolin (Manis pentadactyla) is a critically endangered species. However, there is a paucity of research on the male reproductive gamete biology of this species. The present study was the first to systematically analyse the sperm characterization of the Chinese pangolin, including semen collection, sperm morphometry and ultrastructure. The semen of five male Chinese pangolins was successfully collected using the electroejaculation method. CASA (computer-assisted sperm analysis) was used to assess semen quality and take images for sperm morphometric analysis. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used for sperm ultrastructure observation. The results showed that the semen of the Chinese pangolin was yellow to pale yellow in colour, viscous, with a fishy odour, and a slightly alkaline pH of between 7.7 and 7.9. The head defects were the main sperm defects; there were 13 kinds of head defects counted in this study. The total sperm length, head length, head width and tail length were 67.62 ± 0.21 μm, 10.47 ± 0.06 μm, 1.33 ± 0.006 μm and 57.16 ± 0.20 μm, respectively. SEM observed that the spermatozoa had a rod-shaped head with a distinct apical ridge, which was different from most mammals and similar to that in avians and reptiles. Interestingly, TEM found that the acrosome membrane of the Chinese pangolin had a double membrane structure rather than a multiple bi-lamellar membrane structure as reported by the previous study. Collectively, this study contributes to the development of artificial breeding efforts and assisted reproductive techniques for the Chinese pangolin, as well as providing technical support for research on germplasm conservation of this species.