Determining composition of micron-scale protein deposits in neurodegenerative disease by spatially targeted optical microproteomics.
basic_science · Level V
Where this comes from
- Record sourced from PubMed, PMID 26418743.
- Also identified by DOI 10.7554/eLife.09579 and PMC identifier 4630677.
- Licence recorded as CC BY.
- The licence permits redistribution, so the abstract is shown in full and the full text is available from the publisher.
Abstract
Spatially targeted optical microproteomics (STOMP) is a novel proteomics technique for interrogating micron-scale regions of interest (ROIs) in mammalian tissue, with no requirement for genetic manipulation. Methanol or formalin-fixed specimens are stained with fluorescent dyes or antibodies to visualize ROIs, then soaked in solutions containing the photo-tag: 4-benzoylbenzyl-glycyl-hexahistidine. Confocal imaging along with two photon excitation are used to covalently couple photo-tags to all proteins within each ROI, to a resolution of 0.67 µm in the xy-plane and 1.48 µm axially. After tissue solubilization, photo-tagged proteins are isolated and identified by mass spectrometry. As a test case, we examined amyloid plaques in an Alzheimer's disease (AD) mouse model and a post-mortem AD case, confirming known plaque constituents and discovering new ones. STOMP can be applied to various biological samples including cell lines, primary cell cultures, ex vivo specimens, biopsy samples, and fixed post-mortem tissue.
Medical subject headings
- Mass Spectrometry
- Microscopy, Confocal
- Neurodegenerative Diseases
- Optical Imaging
- Proteins
- Proteomics