Aβ concentration was measured from organoid supernatants using commercially available ELISA kit for Aβ (1–40) and Aβ (1–42) (Life Technologies Corporation, Grand Island, NY) following the manufacturer’s protocol. Briefly, media samples were incubated (4 hours) in primary antibodies against the COOH-terminus of the 1–40 or 1–42 Aβ sequence in pre-coated 96 well plates (pre-coated with monoclonal antibody specific to human Aβ 1–40 or 1–42) followed by aspiration and four washes (in washing buffer) prior to incubation with HRP-conjugated secondary antibodies followed by aspiration, four washes, and addition of HRP substrate (3,3',5,5'-tetramethylbenzidine). The reaction was stopped using 1 N sulfuric acid and absorption was measured at 450 nm in an EnSpire plate readers (Perkin Elmer). Absolute values were calculated from a standard curve and plotted as either picogram/ml (pg/ml) or Aβ42/40 ratio per organoid.
