The profile of soluble amyloid β protein in cultured cell media. Detection and quantification of amyloid β protein and variants by immunoprecipitation-mass spectrometry

Rong Wang, David Sweeney, Samuel E. Gandy, Sangram S. Sisodia

Research output: Contribution to journalArticlepeer-review

290 Scopus citations

Abstract

To study the metabolism of amyloid β protein (Aβ) in Alzheimer's disease, we have developed a new approach for analyzing the profile of soluble Aβ and its variants. In the present method, Aβ and its variants are immunoisolated with Aβ-specific monoclonal antibodies. The identities of the Aβ variants are determined by measuring their molecular masses using matrix- assisted laser desorption ionization time-of-flight mass spectrometry. The levels of Aβ variants are determined by their relative peak intensities in mass spectrometric measurements by comparison with internal standards of known identities and concentrations. We used this method to examine the Aβ species in conditioned media of mouse neuroblastoma cells transfected with cDNAs encoding wild type or mutant human amyloid precursor protein. In addition to human Aβ-(1-40) and Aβ-(1-42), more than 40 different human Aβ variants were identified. Endogenous murine Aβ and its variants were also identified by this approach. The present approach is a new and sensitive method to characterize the profile of soluble Aβ in conditioned media and biological fluids. Furthermore, it allows direct measurement of each individual peptide in a peptide mixture and provides comprehensive information on the identity and concentration of Aβ and Aβ variants.

Original languageEnglish
Pages (from-to)31894-31902
Number of pages9
JournalJournal of Biological Chemistry
Volume271
Issue number50
DOIs
StatePublished - 1996
Externally publishedYes

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