ToF-SIMS lipidomic data for surface lipid content of EcO83-EVs and parent bacteria

For ToF-SIMS measurements we used silicon wafers (cat. No. 647780, Sigma Aldrich, St. Louis, MO, USA) with dimensions of 1 x 1 cm2. Before depositing EVs and cells, each substrate was sonicated in toluene (cat. No. 244511, Sigma Aldrich) and absolute ethanol (99%, cat. No. 396480111, POCH) for 10 min in an ultrasonic bath. We used a ToF-SIMS 5 instrument (ION-ToF GmbH, Münster, Germany) with a Bi3+ liquid metal ion gun (30 keV) as the primary ion source to record data in the mass range up to 900 Da. The experiments were performed in the static spectrometer mode with a current equal to 1.10 pA, and a low-energy electron gun was used to neutralize the charge formed on the sample surface. All mass spectra were measured simultaneously and under the same conditions using a single sample holder.
The normalization of raw ToF-SIMS data is typically performed to eliminate artefacts induced by topographic, apparatus, or matrix effects. To obtain semi-quantitative information from the measured data, normalization was done using the total dose deposited on the examined surface, and each spectrum was calibrated using signals for positive ions: 𝐻+, 𝐻2+, 𝐶𝐻+, 𝐶𝐻2+, 𝐶𝐻3+ and 𝐶3𝐻2+. The obtained spectra were further analyzed, which included the identification of characteristic lipid peaks such as protonated ions, adducts, and pseudo-molecular ions. It should be noted that different molecular formulas may correspond to the same molecular weight.