Importance of manual validation for the identification of phosphopeptides using a linear ion trap mass spectrometer.

ABSTRACT: Accurate determination of protein phosphorylation is challenging, particularly for researchers who lack access to a high-accuracy mass spectrometer. In this study, multiple protocols were used to enrich phosphopeptides, and a rigorous filtering workflow was used to analyze the resulting samples. Phosphopeptides were enriched from cultured rat renal proximal tubule cells using three commonly used protocols and a dual method that combines separate immobilized metal affinity chromatography (IMAC) and titanium dioxide (TiO(2)) chromatography, termed dual IMAC (DIMAC). Phosphopeptides from all four enrichment strategies were analyzed by liquid chromatography-multiple levels of mass spectrometry (LC-MS(n)) neutral-loss scanning using a linear ion trap mass spectrometer. Initially, the resulting

SUBMITTER: Goldstrohm DA 

PROVIDER: S-EPMC3059535 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature