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Peptides were separated using a C18 analytical column NanoEase MZ HSS T3 column (75 μm Å∼ 250 mm, 1.8 μm, 100 Å) (Waters) with a 90 min run, comprising three consecutive steps with linear gradients from 3 to 35% B in 60 min, from 35 to 50% B in 5 min, and from 50 % to 85% B in 2 min, followed by isocratic elution at 85% B in 5 min and stabilization to initial conditions (A= 0.1% formic acid in water, B= 0.1% formic acid in acetonitrile)

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