was added and the tubes and placed on ice for 15 min for dehydration. Before incubation incubated at 37°C for 16 h, 200 µL of 25 mM NH4HCO3 in 50% MeCN was added and the samples. Thereafter, the solution was transferred to clean tubes and 100 µL of 50% MeCN/5% formic acid was added into gel pieces, vortexed for 30 min and the extracted digested pooled together. The in-gel digest protocol was repeated several times, pooled together and the extracted protein digest samples were concentrated.