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Ion mobility-mass spectrometry (IM-MS) provides rapid two-dimensional separations based on analyte apparent surface area or collision cross section (CCS, A(2)) and mass-to-charge, respectively. Recently, traveling-wave (t-wave) IM-MS was developed which uses electrodynamic rather than electrostatic fields commonly used in drift cell IM-MS instruments. The underlying theory for obtaining CCS data is well developed for drift cell IM-MS, while strategies for obtaining CCS values from t-wave IM-MS data remains an active area of research. In this report, methods were developed and validated to obtain CCS values of phospholipids and peptides directly from thin tissue sections by MALDI t-wave IM-MS using CCS calibrants measured by MALDI drift cell IM-MS. Importantly, the average percent difference between t-wave and drift cell CCS measurements is minimized by calibrating with the same biomolecular class. Calibrating t-wave phospholipid CCS values with drift cell peptide CCS measurements results in an average percent difference of ca. 7% between the same lipids measured using t-wave and drift cell IM-MS, while this improves to &lt;0.5% when drift cell phospholipid CCS values are used for calibrating t-wave data. A suite of CCS values are reported for lipids and peptides that were determined directly from tissue, i.e. without the need for tissue extraction and further purification steps.

Structural Characterization of Phospholipids and Peptides Directly from Tissue Sections by MALDI Traveling-Wave Ion Mobility-Mass Spectrometry