Eksigent ekspert nanolc 400 system

LC-MS analysis was performed with AB Sciex Triple ToF 5600+ (AB SCIEX, CA, USA) integrated with LC-MS/MS Eksigent ekspert™ nanoLC 400 System (AB SCIEX, CA, USA). Peptides were separated using nanoACQUITY UPLC 1,8 μM HSS T3 C18 column (Thermo Fisher, MS, USA) in the trap-elute mode. In order to separate the peptides, 4–40% ACN gradient was used for 240 minutes. Data dependent acquisition (DDA) MS/MS analysis of separated peptides was performed after electrospray ionization. Raw data analysis—generated by instrument reporting—and multiple analytical data measurements in each sample were performed with Analyst® TF v.1.6 (AB SCIEX, CA, USA). The peptides and the ion-product of the MS and MS/MS data were evaluated with PeakView (AB SCIEX, CA, USA). Generated peak-lists were evaluated in consideration of the UniProtKB-based reference library of the Mus musculus species on our server with ProteinPilot 4.5 Beta (AB SCIEX, CA, USA).

LC-MS analysis was performed with AB Sciex Triple ToF 5600+ (AB SCIEX, CA, USA) integrated with LC-MS/MS Eksigent ekspert™ nanoLC 400 System (AB SCIEX, CA, USA). Peptides were separated using nanoACQUITY UPLC 1,8 μM HSS T3 C18 column (Thermo Fisher, MS, USA) in the trap-elute mode. In order to separate the peptides, 4–40% ACN gradient was used for 240 min. Data dependent acquisition (DDA) MS/MS analysis of separated peptides was performed after electrospray ionization. Raw data analysis—generated by instrument reporting—and multiple analytical data measurements in each sample were performed with Analyst® TF v.1.6 (AB SCIEX, CA, USA). The peptides and the ion-product of the MS and MS/MS data were evaluated with PeakView (AB SCIEX, CA, USA). Generated peak-lists were evaluated in consideration of the UniProtKB-based reference library of the Homo sapiens species on our server with ProteinPilot 4.5 Beta (AB SCIEX, CA, USA).

Shotgun proteomic analyses have been performed with AB SCIEX TripleTOF^® 5600+ integrated to Eksigent ekspert™ nanoLC 400 System (AB Sciex, USA). Trap column (3 μm, ChromXP C18CL) and nanoAQUITY UPLC^® NanoLC column (1.8-μm HSS T3, 75 μm × 150 mm) were used in the trap-elute mode for the separation of peptides. Within the 310-min analysis, Data Dependent Acquisition Top 20 tandem MS was performed. Raw data analysis and multiple analytical measurements in a single sample were done using Analyst^® TF v.1.6 (AB Sciex, USA) software. Precursor and product ion evaluations were completed with PeakView (1.2, AB Sciex). Generated peak lists containing MS and MS/MS spectra were used to identify proteins, protein isoforms, and their modifications with ProteinPilot 4.5 Beta (AB Sciex, USA). Identification was made using the UniProtKB-based reference library of resistant strain A. baumannii BAA 1710 (UP000002446) and non-resistant strain A. baumannii 17978 (UP000006737). During identifying proteins, the false discovery rate was determined to be 1%, and proteins containing at least two identified peptide fragments were considered correct identification.