title: Spatially-directed protein identification from tissue sections by top-down LC-MS/MS with electron transfer dissociation.
abstract: MALDI imaging mass spectrometry (MALDI-IMS) has become a powerful tool for localizing both small molecules and intact proteins in a wide variety of tissue samples in both normal and diseased states. Identification of imaged signals in MALDI-IMS remains a bottleneck in the analysis and limits the interpretation of underlying biology of tissue specimens. In this work, spatially directed tissue microextraction of intact proteins followed by LC-MS/MS with electron transfer dissociation (ETD) was used to identify proteins from specific locations in three tissue types; ocular lens, brain, and kidney. Detection limits were such that a 1 μL extraction volume was sufficient to deliver proteins to the LC-MS/MS instrumentation with sufficient sensitivity to detect 50-100 proteins in a single experiment. Additionally, multiple modified proteins were identified; including truncated lens proteins that would be difficult to assign to an imaged mass using a bottom-up approach. Protein separation and identification are expected to improve with advances in intact protein fractionation/chromatography and advances in interpretation algorithms leading to increased depth of proteome coverage from distinct tissue locations.
authors.affiliation: Department of Biochemistry and Mass Spectrometry Research Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA. kevin.schey@vanderbilt.edu
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Schey
authors.forename: David M
authors.initials: DM
authors.lastname: Anderson
authors.forename: Kristie L
authors.initials: KL
authors.lastname: Rose
mesh_terms: D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods; D055664:Electrochemical Techniques / Q000379:methods*; D059627:Liquid Phase Microextraction / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods; D051379:Mice; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014020:Tissue Extracts / Q000032:analysis*
chemical_list: D014020:Tissue Extracts