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Kitazume S, Tachida Y, Kato M, Yamaguchi Y, Honda T, Hashimoto Y, Wada Y, Saito T, Iwata N, Saido T, Taniguchi N.
Brain endothelial cells produce amyloid {beta} from amyloid precursor protein 770 and preferentially secrete the O-glycosylated form. J Biol Chem.
2010 Dec 17;285(51):40097-103.
PubMed Abstract
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REAGENTS/MATERIAL:
An anti-APP (C15) rabbit polyclonal antibody against endogenous membrane-bound (intact) APP was a generous gift from Dr. Kei Maruyama (Saitama Medical University, Saitama, Japan) WB, IP.
The commercially available antibodies used were: mouse monoclonal anti-APP (22C11) (Chemicon) WB, IH;
mouse monoclonal anti-human sAPPα (6E10) (Signet Laboratories) WB;
anti-GAPDH (Chemicon) IC, WB;
anti-KPI (Chemicon) WB;
anti-OX2 (Chemicon) WB, IH;
anti-human sAPPβ (IBL Co.) WB, IH and
goat polyclonal anti-PECAM (Santa Cruz Biotechnology) WB.
For Aβ quantification, BMECs were
infected with adenovirus preparations for
APP770-FLAG overexpression, and cultured
in Opti-MEM for 8 h. The levels of Aβ40 and
Aβ42 in the media were determined using a
human Amyloid β(1–40)
or Amyloid β(1–42) Assay Kit
(IBL Co.) for BMECs,
and a Human/Rat
βAmyloid (40) or
βAmyloid (42) ELISA Kit (Wako Chemicals) for mouse neurons.
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