Bovine tuberculosis (bTB) is a chronic infectious disease caused by the Mycobacterium bovis (M. bovis). Rapid, cost-effective, and accurate diagnosis of bTB remains a significant clinical challenge globally. The ideal biomarker for diagnosing bTB would be a molecular derived from M. bovis that can not only confirm the presence of the disease but also help determine its clinical stage. Research has indicated that exosomes, which are small vesicles released by cells, can carry components from pathogen following infection with Mycobacterium tuberculosis (Mtb). In this study, a detailed proteomic analysis was conducted to explore the protein expression profiles in the plasma and plasma exosomes of five cows infected with M. bovis. The findings revealed that exosome samples contained a greater number of M. bovis-derived proteins compared to plasma samples. Specifically, six M. bovis-derived proteins, Mb0416, Mb0704, Mb1051, Mb2900, Mb3904, and Mb3905, were identified in plasma exosomes with high quality spectra. Notably, two of these proteins, Mb3904 and Mb3905, correspond to well-known clinical markers for human tuberculosis, Rv3874 and Rv3875, respectively. Furthermore, exosome host proteins were found to be enriched in more immune-related pathways and biological processes, such as antigen processing and presentation, complement and coagulation cascades, innate immune response, and antimicrobial humoral immune response. These proteins demonstrate immunogenic properties and hold potential as biomarkers for bTB diagnosis, providing new insights for the development of vaccines and therapeutic strategies.