Abstract
The matrix application technique is critical to the success of a matrix-assisted laser desorption/ionization (MALDI) experiment. This work presents a systematic study aiming to evaluate three different matrix application techniques for MALDI mass spectrometric imaging (MSI) of endogenous metabolites from legume plant, Medicago truncatula, root nodules. Airbrush, automatic sprayer, and sublimation matrix application methods were optimized individually for detection of metabolites in the positive ionization mode exploiting the two most widely used MALDI matrices, 2,5-dihydroxybenzoic acid (DHB) and α-cyano-4-hydroxycinnamic acid (CHCA). Analytical reproducibility and analyte diffusion were examined and compared side-by-side for each method. When using DHB, the optimized method developed for the automatic matrix sprayer system resulted in approximately double the number of metabolites detected when compared to sublimation and airbrush. The automatic sprayer method also showed more reproducible results and less analyte diffusion than the airbrush method. Sublimation matrix deposition yielded high spatial resolution and reproducibility but fewer analytes in the higher m/z range (500-1000 m/z). When the samples were placed in a humidity chamber after sublimation, there was enhanced detection of higher mass metabolites but increased analyte diffusion in the lower mass range. When using CHCA, the optimized automatic sprayer method and humidified sublimation method resulted in double the number of metabolites detected compared to standard airbrush method.