Optimized Protocol for On-Target Phosphopeptide Enrichment Prior to Matrix-Assisted Laser Desorption−Ionization Mass Spectrometry Using Mesoporous Titanium Dioxide

摘要

A novel on-target phosphopeptide enrichment method isnpresented that allows specific enrichment and directnanalysis by matrix assisted laser desorption ionizationnmass spectrometry (MALDI-MS) of phosphorylated pep-ntides. Spots consisting of a thin film of anatase titaniumndioxide are sintered onto a conductive glass surface.nEnrichment and analysis can be performed on the modi-nfied target with minimal sample handling. The protocolnleads to an enrichment efficiency that is superior to whatnhas been reported before for similar methods. The methodnwas tested using u0001-casein as a model phosphorylatednprotein as well as with a custom peptide mixed with itsnphosphorylated form. A very low detection limit, a signifi-ncantly improved phosphoprofiling capability, and a simplenexperimental approach provide a powerful tool for thenenrichment, detection, and analysis of phosphopeptides.