Spatially-Resolved MALDI-MS Imaging of Metabolites

Muscle tissues were sectioned at 10-μm thickness with a cryostat and thaw-mounted onto ITO-coated glass slides. 9-AA (600 mg) was deposited on slides at 0.5-μm thickness in an iMLayer (Shimadzu, Kyoto, Japan), and recrystallization was carried out by the methods described previously with slight modification47 (link). In this study, Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometer (MALDI-TOFMS, AXIMA^® Confidence, Shimadzu) equipped with a 337 nm N₂ laser was used for mass spectrometry analysis. Mass spectra were acquired with the laser frequency in the negative and scanning mass range from m/z 50 to m/z 1,000 at a high-resolution mode. Laser power, detection voltage, and accumulated number of MALDI-TOFMS were 115, 3.0 kV, and 1/pixel, respectively. The spatial interval of data points was 50 μm, giving 14641 profiles in total for the section. Data collected through the microscopic system were digitally processed using Imaging MS Solution analysis software (Shimadzu). The signal intensity of each imaging data in the figure is represented as the normalized intensity. Metabolites were identified with the MS/MS spectrum from results of IS chemical standard analysis using MALDI-QIT-TOFMS (AXIMA^® QIT, Shimadzu).