Representative diagram of the MALDI-MSI strategy. (A) The epidermis of whole leaves can be sprayed with a matrix directly or the tissue is cut into 10-15 μm sections and the sections are coated with a matrix on glass slides. The data are acquired across the section in the form of a raster of predefined resolution, and a mass spectrum of each raster-point is recorded. For each m/z value the ion intensity and distribution is collected. (B) Two options for identification of potential biomarkers are proposed. In some case, the slide may be coated with trypsin for on-slide protein digestion and peptides are analysed as above using reflectron mode and MS/MS. More commonly, the tissue region of interest needs to be excised and separated, isolating the m/z ion of interest. The ion of interest can then be digested with trypsin and analysed using LC-MS/MS.