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Mass Spectrometry Imaging of Over 2,000 Proteins from Tissue Sections at Better than 100-µm Spatial Resolution using nanoPOTS Sample Preparation
Monday, March 08, 2021: 1:30 PM - 2:05 PM
Speaker(s)
Description
Biological tissues contain a wide variety of cell types with distinct functions and understanding this heterogeneity at high spatial resolution is of great interest in biomedical research. Immunohistochemistry staining methods are limited to visualizing fewer than 10 protein targets, and the image quality is limited by the specificity of antibodies. Imaging Mass spectrometry (IMS) provides a label-free approach to directly visualize peptides and proteins from tissue sections, however technical challenges have limited proteome coverage of IMS to the most abundant proteins. Further, the effects of the tissue matrix on ionization efficiency of biomolecules can greatly diminish the quantification accuracy. To address these challenges, we developed an integrated IMS platform that combines laser capture microdissection, nanoPOTS (nanodroplet Processing in One pot for Trace Samples) sample preparation, nanoLC-MS/MS, and an open-source bioinformatic tool (Trelliscope) for data processing and visualization. We demonstrated that the nanoPOTS IMS platform can quantitatively generate cell-type-specific images for >2,000 proteins at 100-µm spatial resolution across 12-µm thick tissue sections using a label free quantitation approach. We have also demonstrated that incorporating tandem mass tags (TMT) labeling with our workflow can improved analysis throughput, overall sensitivity, and enabled MS imaging at 50-µm spatial resolution, without compromising proteome coverage. The ability of this nanoPOTS imaging platform to map quantitative protein changes across cell-types and cellular microenvironments in mouse and human thin tissue sections demonstrates its potential to advance biomedical research. This work was supported by the National Institutes of Health (NIH) NICHD under award R21 HD084788 and NIH Common Fund, through the Office of Strategic Coordination/Office of the NIH Director under award UG3HL145593 (HuBMAP Program).
Additional Info
Keywords: Please select up to 4 keywords ONLY:
Automation,Biomedical,Proteomics,Nanoscience
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