title: Microextraction and purification techniques applicable to chemical characterization of amyloid proteins in minute amounts of tissue.
abstract: This article described micromethods useful for the extraction, purification, and amino acid sequencing of amyloid proteins contained in minute specimens obtained from patients with systemic forms of amyloidosis. We posit that these procedures can also be applied to the biochemical characterization of cerebral amyloid deposits. The selection of the techniques is dependent on the type of sample to be extracted (fresh or formalin fixed) as well as the amount of congophilic material present. Although amyloid proteins are isolated and purified more easily from fresh tissue, it must be noted that formalin-fixed specimens are available more readily for analysis due to the common diagnostic use of fine needle tissue biopsies and are therefore, important for both current and retrospective studies. Remarkably, despite the expected difficulties associated with formalin treatment we were able to extract and sequence amyloid proteins from fixed tissues presumably due to the resistance of amyloid to formalin cross-linking. Through the continued development of techniques for small-scale protein separation and application of highly sensitive microsequencing and mass spectral methods, exact identification of the protein contained in fibrillar amyloid deposits can be determined. Such information has therapeutic and prognostic relevance and can increase our understanding of the pathogenesis of amyloidosis.
authors.affiliation: Heller Institute of Medical Research, Chaim Sheba Medical Center, Tel Hashomer, Israel.
authors.forename: B
authors.initials: B
authors.lastname: Kaplan
authors.forename: R
authors.initials: R
authors.lastname: Hrncic
authors.forename: C L
authors.initials: CL
authors.lastname: Murphy
authors.forename: G
authors.initials: G
authors.lastname: Gallo
authors.forename: D T
authors.initials: DT
authors.lastname: Weiss
authors.forename: A
authors.initials: A
authors.lastname: Solomon
mesh_terms: D000544:Alzheimer Disease / Q000378:metabolism; D000595:Amino Acid Sequence; D000682:Amyloid / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006801:Humans; D008969:Molecular Sequence Data; D017421:Sequence Analysis / Q000379:methods*
chemical_list: D000682:Amyloid