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Second-dimension separation was performed as described by OFarrell (1975) in large format (20 cm 20 cm), 1.5 mm thick 12% SDS-PAGE gels in standard Tris-glycine buffer in a PROTEAN II xi cell for vertical electrophoresis (BioRad, United States)

Before mass spectrometry, the supernatant should be redissolved in 100 L of acetonitrile solution (1:1 ratio), vortexed, and centrifuged again at 14 000 g at 4C for 15 min to prepare for analysis

in 1968 [41]

Once reconstituted into a liquid solution, degradation begins, and the stability window shortens significantly

The plateau emerges when another bottleneck becomes dominant