This project explores the proteomic alterations associated with polyhexamethylene biguanide (PHMB) resistance in Staphylococcus aureus using tandem mass tag (TMT)-based quantitative proteomics. PHMB-resistant and control strains were compared to identify proteins involved in resistance mechanisms, particularly those related to cell envelope modification and surface charge regulation. Proteins encoded by the dlt operon were found to be significantly upregulated in the resistant strain, suggesting their involvement in PHMB tolerance and potential cross-resistance to other antibiotics. This dataset provides comprehensive proteomic evidence for understanding the molecular basis of PHMB-induced resistance in S. aureus.