title: Intraneuronal Aβ detection in 5xFAD mice by a new Aβ-specific antibody.
abstract: BACKGROUND
  The form(s) of amyloid-β peptide (Aβ) associated with the pathology characteristic of Alzheimer's disease (AD) remains unclear. In particular, the neurotoxicity of intraneuronal Aβ accumulation is an issue of considerable controversy; even the existence of Aβ deposits within neurons has recently been challenged by Winton and co-workers. These authors purport that it is actually intraneuronal APP that is being detected by antibodies thought to be specific for Aβ. To further address this issue, an anti-Aβ antibody was developed (MOAB-2) that specifically detects Aβ, but not APP. This antibody allows for the further evaluation of the early accumulation of intraneuronal Aβ in transgenic mice with increased levels of human Aβ in 5xFAD and 3xTg mice.
  
  
  RESULTS
  MOAB-2 (mouse IgG2b) is a pan-specific, high-titer antibody to Aβ residues 1-4 as demonstrated by biochemical and immunohistochemical analyses (IHC), particularly compared to 6E10 (a commonly used commercial antibody to Aβ residues 3-8). MOAB-2 did not detect APP or APP-CTFs in cell culture media/lysates (HEK-APPSwe or HEK-APPSwe/BACE1) or in brain homogenates from transgenic mice expressing 5 familial AD (FAD) mutation (5xFAD mice). Using IHC on 5xFAD brain tissue, MOAB-2 immunoreactivity co-localized with C-terminal antibodies specific for Aβ40 and Aβ42. MOAB-2 did not co-localize with either N- or C-terminal antibodies to APP. In addition, no MOAB-2-immunoreactivity was observed in the brains of 5xFAD/BACE-/- mice, although significant amounts of APP were detected by N- and C-terminal antibodies to APP, as well as by 6E10. In both 5xFAD and 3xTg mouse brain tissue, MOAB-2 co-localized with cathepsin-D, a marker for acidic organelles, further evidence for intraneuronal Aβ, distinct from Aβ associated with the cell membrane. MOAB-2 demonstrated strong intraneuronal and extra-cellular immunoreactivity in 5xFAD and 3xTg mouse brain tissues.
  
  
  CONCLUSIONS
  Both intraneuronal Aβ accumulation and extracellular Aβ deposition was demonstrated in 5xFAD mice and 3xTg mice with MOAB-2, an antibody that will help differentiate intracellular Aβ from APP. However, further investigation is required to determine whether a molecular mechanism links the presence of intraneuronal Aβ with neurotoxicity. As well, understanding the relevance of these observations to human AD patients is critical.
authors.affiliation: Department of Anatomy and Cell Biology, University of Illinois at Chicago, IL 60612, USA.
authors.forename: Katherine L
authors.initials: KL
authors.lastname: Youmans
authors.forename: Leon M
authors.initials: LM
authors.lastname: Tai
authors.forename: Takahisa
authors.initials: T
authors.lastname: Kanekiyo
authors.forename: W Blaine
authors.initials: WB
authors.lastname: Stine
authors.forename: Sara-Claude
authors.initials: SC
authors.lastname: Michon
authors.forename: Evelyn
authors.initials: E
authors.lastname: Nwabuisi-Heath
authors.forename: Arlene M
authors.initials: AM
authors.lastname: Manelli
authors.forename: Yifan
authors.initials: Y
authors.lastname: Fu
authors.forename: Sean
authors.initials: S
authors.lastname: Riordan
authors.forename: William A
authors.initials: WA
authors.lastname: Eimer
authors.forename: Lester
authors.initials: L
authors.lastname: Binder
authors.forename: Guojun
authors.initials: G
authors.lastname: Bu
authors.forename: Chunjiang
authors.initials: C
authors.lastname: Yu
authors.forename: Dean M
authors.initials: DM
authors.lastname: Hartley
authors.forename: Mary Jo
authors.initials: MJ
authors.lastname: LaDu
mesh_terms: D000544:Alzheimer Disease / Q000473:pathology; D016229:Amyloid beta-Peptides / Q000032:analysis* / Q000276:immunology; D016564:Amyloid beta-Protein Precursor / Q000032:analysis* / Q000276:immunology; D000818:Animals; D000906:Antibodies / Q000032:analysis* / Q000276:immunology; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D002478:Cells, Cultured; D004195:Disease Models, Animal; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D058225:Plaque, Amyloid / Q000737:chemistry / Q000473:pathology
chemical_list: D016229:Amyloid beta-Peptides; D016564:Amyloid beta-Protein Precursor; D000906:Antibodies