Cook JJ, Wildsmith KR, Gilberto DB, Holahan MA, Kinney GG, Mathers PD, Michener MS, Price EA, Shearman MS, Simon AJ, Wang JX, Wu G, Yarasheski KE, Bateman RJ. Acute gamma-secretase inhibition of nonhuman primate CNS shifts amyloid precursor protein (APP) metabolism from amyloid-beta production to alternative APP fragments without amyloid-beta rebound. J Neurosci. 2010 May 12;30(19):6743-50. PubMed Abstract, View on AlzSWAN

REAGENTS/MATERIAL:
Amyloid β ELISA: Solid black 96 well Nunc plates were coated with 3 μg/ml N-terminal mouse monoclonal anti-Aβ (6E10) (Signet) in pH 9.4 bicarbonate buffer overnight at 4°C. CSF samples for the Aβ1-40 assay were diluted 1:8 in BSA/PBS; 50 μl of samples and standards, starting at 4:00 P.M., were loaded onto the plate. CSF samples for Aβ1-42 were diluted 1:4 in BSA/PBS and 100 μl of samples and standards, starting at 2:00 P.M., were loaded onto the plate. Plasma samples were diluted 1:4 in BSA/PBS and 50μl of samples and standards, starting at 4:00 P.M., were loaded onto the plate. Fifty microliters of G2-10-alkaline phosphatase (Aβ1-40) or 12F4-alkaline phosphatase (Aβ1-42) diluted in BSA/PBS was added to the plate and incubated overnight at 4°C.
Amyloid β immunoprecipitation: Antibody beads were prepared by covalently binding 4G8 antibody (GE Healthcare) (recognizes Aβ residues 18-22) or anti-Aβ 6E10 antibody (Covance) (recognizes Aβ residues 1-17) to CNBr Sepharose beads (GE Healthcare).