The engineered Staphylococcus aureus strain RN4220Δagr/EDⅢ was treated with 1/4 MIC of Oxacillin (OXA), or used to construct pbp3 and pbp4 gene mutant strain. Then, the membrane vesicles (MVs) of parent strain (ΔagrMVs), OXA-treated strain (Δagr/OXAMVs), and pbp3/pbp4 mutants (Δagr/OXAMVs) were harvested by density gradient centrifugation. The protein compositions of each MV sample were quantitated by using BCA Protein Assay Kit.