phosphoproteome

Phospho-peptide enrichment Protein extraction, reduction, alkylation and digestion were conducted as described above. Then, 15 mg TiO2-coupled beads were incubated with 500 μl Binding Buffer (BB) at RT for 10 min. Each aliquoted TiO2 was added into three 1.5 ml EP tubes, 5 mg for each and centrifuge 2,000 g for 2 min. The peptides were re-solved with 6μl BB and combined with 5 mg incubated TiO2 at RT for 30 min and centrifuge 1,000 g for 2 min. Repeat the solved procedures for the dried phosphopeptides twice with the 5 mg TiO2 and then discard the supernatant (Tong et al., 2019a).