Amazon sl quadrupole ion trap mass spectrometer

Following desalting with C18 columns, the peptides were eluted, dried and re-suspended in 0.1% formic acid (FA) then subjected to LC-MS/MS using an Agilent 1200 series micro flow HPLC coupled to a Bruker Amazon-SL quadrupole ion trap mass spectrometer, and a captive spray ionization source. Tryptic peptides were separated at a solvent flow rate of 1.6 μL/min with 0 to 40% gradients of 0.1% FA (solvent A) and acetonitrile (ACN) in 0.1% FA (solvent B) over a period of 300 min each using a C₁₈ (150 × 0.1 mm, 3.5 μm particle size, 300 Å pore size, Zorbax SB) capillary column. The captive electrospray source was operated in a positive ion mode with a dry gas temperature of 150 °C, dry nitrogen flow 3 L/minute, and capillary voltage of 1500 V. The data were acquired in the auto MS (n) mode with optimized trapping condition for the ions at m/z 1000. MS scans were performed in the enhanced scanning mode (8100 m/z/second), while the collision-induced dissociation or the MS/MS fragmentation scans performed automatically for top ten precursor ions for 1 min each in the UltraScan mode (32,500 m/z/second). The samples were run three times as technical repeats.

Lipids were extracted from bacterial cultures as described elsewhere^{62 (link)}. Briefly, bacteria were grown to OD₆₀₀ of 1.0 in TSB or TSB supplemented with 10 µM LA, centrifuged in a 2 mL glass Chromacol vial (Thermo Scientific), and resuspended in 0.5 mL MS grade methanol (Sigma-Aldrich). MS grade chloroform was then used to extract lipids. The extracted lipids were dried under nitrogen gas with a Techne sample concentrator (Staffordshire, UK), and the lipid pellets resuspended in 1 mL acetonitrile. The samples were then analysed by LC–MS with a Dionex 3400RS HPLC coupled to an amaZon SL quadrupole ion trap mass spectrometer (Bruker Scientific) via an electrospray ionisation interface. Both positive and negative ionisation modes were used for sample analyses. The Bruker Compass software package was utilized for data analyses, using DataAnalysis for peak identification and characterization of lipid class, and QuantAnalysis for quantification of the relative abundance of distinct PG species to total PG species.