Automated platform of μLC-MS/MS using SAX trap column for highly efficient phosphopeptide analysis.

In the current study, a specially designed automated nanoflow liquid chromatography coupled with tandem mass spectrometer (μLC-MS/MS) system for phosphopeptide analysis was reported. The system was established by applying a strong anion exchange (SAX) trap column. At the beginning of the analysis, phosphopeptides were loaded onto SAX trap column at high flow rate. Next, the retained phosphopeptides were eluted onto a C18 analytical capillary column by an acidic buffer with high ionic strength. At last, its performance was evaluated by analyzing the phosphopeptides enriched from the tryptic digest of mouse liver lysate. Compared with conventional automated μLC-MS/MS system using C18 trap column, much more phosphopeptides could be identified by the SAX trap column system. This system was fully automated and had promising application in high throughput phosphoproteomic analysis.