Matrix Application for MALDI-IMS

The MALDI matrix 2,5-dihydroxybenzoic acid (DHB) (Sigma-Aldrich, St. Louis, MO, USA) was prepared in 60:40 ACN/H₂O with 0.2% TFA (EMD, Billerica, MA, USA) to a final concentration of 30 mg/mL. Before use, the matrix solution was filtered (0.22 μm filter) and applied to samples using a TM-Sprayer (HTX Technologies, Carrboro, NC, USA) with heated nitrogen sheath flow gas. The solvent pump flow rate was set to 0.1 mL/min and the temperature was set to 70 °C. Matrix was applied five times in a crisscross pattern; samples were dried in a desiccator for 1 h prior to MALDI-MSI.