title: Sensitive, Simple, and Robust Nano-Liquid Chromatography-Mass Spectrometry Method for Amyloid Protein Subtyping.
abstract: Amyloidosis is a rare condition characterized by deposits of insoluble proteins in the form of β-pleated sheets. These deposits interfere with the normal structure and function of varying tissues. Thirty-one amyloid proteins have been identified, and the correct identification is critical due to the varying treatments. Immunohistochemistry, the most routine method for identification of amyloid proteins, suffers from limitations. Mass spectrometry (MS)-based methods offer better sensitivity and specificity. We describe here a sensitive, simple, and robust MS-based method for the identification of amyloid proteins. Amyloid deposits are excised from formalin-fixed tissue by laser microdissection and is put through protein extraction followed by trypsin digestion. The resulting peptides are separated by nano-liquid chromatography and analyzed by high-resolution Orbitrap mass spectrometry. The mass spectrometry data are then searched against a human protein database for identification and semi-quantification.
authors.affiliation: Department of Clinical Pathology, Cleveland Clinic, Cleveland, OH, USA.
authors.forename: Drew
authors.initials: D
authors.lastname: Payto
authors.affiliation: Department of Clinical Pathology, Cleveland Clinic, Cleveland, OH, USA.
authors.forename: Courtney
authors.initials: C
authors.lastname: Heideloff
authors.affiliation: Department of Laboratory Medicine, Cleveland Clinic, Cleveland, OH, USA. wangs2@ccf.org.
authors.forename: Sihe
authors.initials: S
authors.lastname: Wang
mesh_terms: D058227:Amyloidogenic Proteins / Q000378:metabolism*; D002853:Chromatography, Liquid / Q000379:methods*; D030562:Databases, Protein; D006801:Humans; D057230:Limit of Detection; D013058:Mass Spectrometry / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D040901:Proteomics / Q000379:methods*
chemical_list: D058227:Amyloidogenic Proteins
keywords: Amyloidogenic protein; Amyloidosis; Immunohistochemistry; Orbitrap mass spectrometry; β-pleated sheets