Sample preparation for mass spectrometry imaging of leaf tissues: a case study on analyte delocalization

Appropriate sample preparation is pivotally important to obtain high-quality mass spectrometry imaging (MSI) data. Unlike mammalian tissues, preparation of cryosections from plant tissues for MSI measurement is quite challenging due to its intrinsic complex texture and cellular structure. This is especially true for leaf samples which are generally thin, water-rich, and fragile. In this work, a systematic study was performed, aiming to evaluate three embedding materials and five mounting approaches for matrix-assisted laser desorption ionization (MALDI) MSI of secondary metabolites in cross sections of the ginkgo leaf. Delocalization of endogenous metabolites was chosen as a major indicator for evaluation of three embedding materials including ice, carboxymethyl cellulose (CMC), and gelatin and different mounting approaches. Image distortion and analyte delocalization were observed when ice was used as an embedding medium. CMC embedding provided better results compared to the ice by using modified mounting approach. Among three embedding materials, no delocalization was observed in specimens embedded with gelatin, and gelatin embedding is the least affected by different mounting approaches. An alternative approach to mitigate analyte delocalization is the removal of embedding media embraced the tissue sections before mounting, which is particularly suitable for ice-embedded samples. Additionally, the extent of analyte delocalization was closely related to their lipophilicity/hydrophilicity properties, and less analyte diffusion was observed for hydrophobic analytes than for the water-soluble compounds.