Matrix-Assisted Laser Desorption Ionization Mass Spectrometry Imaging by Freeze-Spot Deposition of the Matrix

Imaging mass spectrometry has emerged as a powerful metabolite measurement approach to capture the spatial dimension of metabolite distribution in a biological sample. In matrix- assisted laser desorption ionization-mass spectrometry imaging (MALDI-MSI), deposition of the chemical-matrix onto the sample serves to simultaneously extract biomolecules to the sample surface and concurrently render the sample amenable to MALDI. However, matrix application may mobilize sample metabolites and will dictate the efficiency of matrix crystallization, together limiting the lateral resolution which may be optimally achieved by MSI. Here, we describe a matrix application technique, herein referred to as the "freeze-spot" method, conceived as a low-cost preparative approach requiring minimal amounts of chemical matrix while maintaining the spatial dimension of sample metabolites for MALDI-MSI. Matrix deposition was achieved by pipette spot application of the matrix-solubilized within a solvent solution with a freezing point above that of a chilled sample stage to which the sample section is mounted. The matrix solution freezes on contact with the sample and the solvent is removed by sublimation, leaving a fine crystalline matrix on the sample surface. Freeze-spotting is quick to perform, found particularly useful for MALDI-MSI of small sample sections, and well suited to efficient and cost-effective method development pipelines, while capable of maintaining the lateral resolution required by MSI.