Site-to-site reproducibility of Matrix-Assisted Laser Desorption Ionization Mass Spectrometry Imaging from Formalin-Fixed Paraffin-Embedded samples

Background & Objective Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI-MSI) allows extraction of molecular information from tissue sections and can be used to create clinically relevant classification models, also known as tissue typing. For widespread clinical application, MALDI-MSI must analyze Formalin-Fixed Paraffin-Embedded (FFPE) tissues. Our aim was to determine if MALDI-MSI of FFPE samples is reproducible when obtained from different sites. Methods All sites used a standard operating protocol (SOP) which also covered instruments. FFPE tissues (3-5µm; mouse intestine, tissue microarray of tumor entities sampled from three different sites) were prepared for MALDI-MSI. Samples were coated with trypsin using an automated sprayer then incubated in a humid environment. After digestion, alpha-cyano-4-hydroxycinnamic acid was deposited using the same sprayer and analyzed with a rapifleX MALDI Tissuetyper. After acquisition, statistical analysis and segmentation feature extraction was conducted. Results Mouse intestine was used to assess whether operators performing an idealized experiment at different sites can obtain similar results, while a TMA consisting of different tumor samples was used to investigate if different tissue sampling sites introduces variation. Eight measurements of the mouse intestine split over five sites and two time points showed reproducible delineation of the villi from underlying muscle. Measurements of the TMA indicated that tissue biology has a greater influence on the spectra than location of sample origin. Conclusion Preliminary results indicate that strict adherence to a SOP produces reproducible MALDI-MSI data from FFPE samples and that sampling site is not a major source of variation.