Immunoblotting and Tandem Mass Spectrometry Analyses

This experiment refers to the previous research method (Huang et al., 2016a). A. baumannii Ab112 whole cells (WCs) and OMVs were separated by SDS-PAGE. WCs were transferred to a PVDF membrane. A. baumannii Ab112 OMV (AbOMV) antiserum or control serum was used as the primary antibody (diluted 1:1000), and horseradish peroxidase (HRP)-conjugated anti-mouse IgG (Invitrogen) was used as the secondary antibody at a 1:10,000 dilution. The blots were developed with an electrochemiluminescence (ECL) substrate (Thermo Scientific).

Tandem matrix-assisted laser desorption/Ionization (MALDI)-time of flight (TOF)-TOF mass spectrometry analysis was performed by Sangon Biotech (Shanghai) Co., Ltd., to identify the protein bands of interest by SDS-PAGE. The samples were removed directly from the gel.