Assays

The neurons were lysed direct on the plate in RIPA lysis buffer using a cell scraper. The lysate was transferred into a fresh Eppendorf tube and undissolved material was removed by centrifugation for 5 min at 20,000 g and 4°C. A protein assay was performed and 15 µg of protein were subjected to proteolytic digestion with the SP3 protocol. Proteins were reduced by addition of 9 µL of 200 mM dithiothreitol (Biozol, Germany) in 50 mM ammonium bicarbonate and incubation for 30 min at 37°C. Cysteine ...