Thermo scientific easy nlc 1000

LC-MS/MS analysis was performed using a High Field Q Exactive mass spectrometer coupled to a Thermo Scientific EASY-nLC 1000 (Thermo Fisher Scientific, Waltham, MA) equipped with a self-packed 75 µm x 20-cm reverse phase column (ReproSil-Pur C18, 3 µm, Dr. Maisch GmbH, Germany) for peptide separation. Analytical column temperature was maintained at 50 °C by a column oven (Sonation GmBH, Germany). Peptides were eluted with a 3–40% acetonitrile gradient over 60 min at a flow rate of 250 nL min⁻¹. The mass spectrometer was operated in data-dependent (DDA) mode with survey scans acquired at a resolution of 120,000 over a scan range of 300-1750 m/z. Up to fifteen most abundant precursors from the survey scan were selected with an isolation window of 1.6 Th and fragmented by higher-energy collisional dissociation with Normalised Collision Energies (NCE) of 27. Maximum ion injection time for the survey and MS/MS scans was 60 ms and the ion target value for both scan modes was set to 3e6.

LC-MS/MS analysis was performed using a Q Exactive mass spectrometer coupled to a Thermo Scientific EASY-nLC 1000 (Thermo Fisher Scientific, Waltham, MA) equipped with a self-packed 75 μm × 20-cm reverse phase column (ReproSil-Pur C18, 3 μm, Dr. Maisch GmbH, Germany) for peptide separation. Analytical column temperature was maintained at 50 ^oC by a column oven (Sonation GmBH, Germany). Peptides were eluted with a 3–40% acetonitrile gradient over 60 min at a flow rate of 250 nL min⁻¹. The mass spectrometer was operated in data-dependent (DDA) mode with survey scans acquired at a resolution of 120,000 over a scan range of 300–1750 m/z. Up to fifteen most abundant precursors from the survey scan were selected with an isolation window of 1.6 Th and fragmented by higher-energy collisional dissociation with Normalized Collision Energies (NCE) of 27. Maximum ion injection time for the survey and MS/MS scans was 60 ms and the ion target value for both scan modes was set to 3e6.