Measuring Aβ40 and Aβ42 in hiPSC- and miPSC-derived Neuron Culture Medium

Medium from neuronal cultures was harvested and stored at –80°C. Cell lysates were collected for cellular protein normalization and analysis of p-tau levels by western blot analysis. Human and mouse Aβ peptides were measured with MSD Human V-PLEX Aβ Peptide Panel 1 (6E10) Kit (K15200E, Meso Scale Discovery) and Thermo Fisher Mouse Aβ Peptide ELISA Kits (KMB3481 and KMB3441, Thermo Fisher Scientific) according to the product manuals. For human Aβ measurement, the plates were read with a Sector Imager 2400, and the data were acquired and analyzed with Discovery Workbench software. For mouse Aβ measurement, the plates were read with a FlexStation-III, and the data were analyzed with Prism-6 software. For experiments with secretase inhibitors, neuronal cultures were treated with a γ-secretase inhibitor (compound E, Copd-E, at a final concentration of 200 nM), a β-secretase inhibitor (OM99-2, final concentration of 750 nM), or dimethyl sulfoxide (vehicle) as described¹⁰. All inhibitors were from EMD Millipore.