For phosphoproteome studies, we recommend starting with >1mg amounts of complex protein material. The titanium dioxide enrichment step is used to pull the phopshopeptides away from the non-phosphopeptides and thereby enrich this population of peptides. Both the enriched and un-enriched peptides are analyzed by LC-MS/MS separately. Strong cation fractionation can also be performed as in the Protein-ID-solution-fractions (MudPIT) service.
Suitable Protein Complexity
|PTM-phosphoproteome-solution||Complex protein mixtures with no prior fractionation||Sample clean-up , dual enzymatic digestion, TiO2 phosphopeptide enrichment, and LC-MS/MS on both the enriched and un-enriched fractions (2 LC-MS/MS runs) with a long length gradient, and a database search|
|PTM-phosphoproteome-fractions||More than 1000 proteins with prior fractionation and LC-MS/MS analysis of 10 fractions||Same as above but with fractionation prior to LC-MS/MS (Short length gradient) of 10 fractions. ONLY the enriched fractions are run in the grouped cost.|
- Application Note 6: Using MD-Score and PhosphoRS in YPED to aid in localization and verification of phosphosites
- Sample Submission – YPED