Unless otherwise indicated, all steps below were performed at 4 ℃.
For the extraction of proteins, the sample powder in the Eppendorf tube was mixed with an ice-cold solution containing 5 mM/L TriseHCl, pH 7.5 and 1 mM/L CaCl2, as described by Finnie, Melchior, Roepstorff, and Svensson (2002).