Background: Thymoma is often accompanied by myasthenia gravis (MG), and the resection of thymoma improves myasthenic symptoms in patients with thymoma and MG (TMG), but some patients still have no relief. Through proteomic analysis, we examined preoperative serum samples from patients with TMG to identify key prognostic proteins that could serve as a foundation for clinically predicting postoperative efficacy and guiding treatment selection. Method: In this study, 20 patients with TMG meeting the inclusion criteria were selected and divided into an effective group (T1), an ineffective group (T2) with 10 cases each, and a healthy control group (C) with 9 cases. Blood samples from the three groups were collected through Data independent acquisition (DIA) proteomic analysis performed by mass spectrometry to identify differential proteins expressed and search for key proteins associated with myasthenia prognosis. Finally, the target proteins were validated through the utilization of Enzyme-Linked ImmunoSorbent Assay (ELISA). Results: 514 proteins were identified in this research. Between the T1 and T2 groups, there were 20 proteins that exhibited differential expression, with 10 showing up-regulation and 10 displaying down-regulation. KEGG functional annotation indicated that these proteins were mainly involved in signaling pathways such as complement and coagulation cascade, prion disease, systemic lupus erythematosus, neutrophil extracellular trap formation, and transcription dysregulation in cancer. 3 proteins were discovered to have a significant correlation with the prognosis of TMG, L-selectin (SELL) was down-regulated, HLA class I histocompatibility antigen (HLA-A) and Complement 5 (C5) were up-regulated. ELISA results confirmed the proteomic results. Conclusion: HLA-A, C5 and SELL may be potential prognostic biomarkers of TMG.