TMT labeling 800 µg TMT mass tag reagents were dissolved 80 µl 100% (v/v) anhydrous acetonitrile to obtain final concentrations of 10 µg/µl. Resuspended TMT reagents were incubated at RT for 10 minutes, then vortexed and centrifuged briefly (2000 g / 2 minutes / RT). 50 µg lyophilized peptides were resuspended in 50 µl of a mixture containing 42 µl 50 mM TEAB and 8 µl 100% (v/v) anhydrous acetonitrile. Resuspended peptides were sonicated for 10 minutes, then centrifuged at 17,000 g for 10 minutes at RT. Peptides were transferred to fresh protein low-bind 1.5 ml Eppendorf tubes. 20 µl of 10 µg/µl TMT reagent were added to solubilized peptides, vortexed, centrifuged briefly (2,000 g / 1 minute / RT) and incubated on a Thermomixer for 2 hours at 800 rpm at RT. 50 µl of 50 mM TEAB was added to each reaction, followed by vortex, brief centrifugation (2,000 g / 1 minute / RT) and incubation on a Thermomixer at 800 rpm at RT for an additional 10 minutes. 5 µl of each TMT labeled sample was set aside, vacuum dried and injected on MS to confirm that labeling efficiency was >98%. The remaining reactions were stored in -80°C until labeling efficiency was verified. TMT samples were thawed to RT and labeling reactions were quenched with the addition of 5 µl 5% (v/v) hydroxylamine (dissolved in water from a 50% (v/v) stock solution). Samples were incubated on a Thermomixer for 20 minutes at 800 rpm at RT. Quenched TMT labeled samples were pooled, vacuum dried and subjected to High-pH fractionation as described previously , 96 fractions were collected and were concatenated into 48 fraction. Pooled fractions were vacuum dried and stored in -20 freezer until the LC-MS/MS analysis.