Samples were analyzed by 1D-LC-MS/MS. For each mussel sample, three technical replicates were run. Two wash runs and one blank run were done between samples to reduce carry over. For each run, 2 to 7 µg of peptide were loaded onto a 5 mm, 300 µm ID C18 Acclaim® PepMap100 pre-column (Thermo Fisher Scientific) using an UltiMateTM 3000 RSLCnano Liquid Chromatograph (Thermo Fisher Scientific), and desalted on the pre-column. After desalting the peptides, the pre-column was switched in line with a 50 cm x 75 µm analytical EASY-Spray column packed with PepMap RSLC C18, 2µm material (Thermo Fisher Scientific), which was heated to 45° C. The analytical column was connected via an Easy-Spray source to a Q Exactive Plus hybrid quadrupole-Orbitrap mass spectrometer (Thermo Fisher Scientific). Peptides were separated on the analytical column and mass spectra acquired in the Orbitrap as described previously70 (link). Roughly 650,000 MS/MS spectra were acquired per sample (three technical replicates combined).
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Mussel Proteome Analysis by 1D-LC-MS/MS
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Corresponding Organization : Max Planck Institute for Marine Microbiology
Other organizations : University of Bremen, University of Calgary, University of North Carolina at Charlotte, University of California, Santa Barbara
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Variable analysis
independent variables
- None explicitly mentioned
- MS/MS spectra acquired per sample (three technical replicates combined)
- Wash runs and blank run between samples to reduce carry over
- Pre-column (5 mm, 300 µm ID C18 Acclaim® PepMap100) for desalting peptides
- Analytical column (50 cm x 75 µm C18, 2µm) heated to 45° C
- Connection to Q Exactive Plus hybrid quadrupole-Orbitrap mass spectrometer
- None explicitly mentioned
- Blank run between samples
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