After centrifugation
at 12 000 × g for 15 min at 4°C, the proteins in the supernatant
were precipitated by adding four fold volume
of ice-cold acetone containing 10% TCA and 0.07% (w/v)
β-ME, incubated at -20°C for at least 3 h, and then
centrifuged for 15 min at 12 000 × g.