Human HeLa cells LC MS/MSLC-MSMS

We developed a reagent of 10-plex isobaric tags (IBT) with high stability and low cost. The labeled peptides were sensitively detected on Orbitrap Q Exactive MS with MS2 resolution of 35000 at 30% NCE, while the peptides were efficiently labelled over 97% by IBT at ratio of 20:1 of reagent/peptide (w/w) in 200mM TEAB buffer within 2h. The IBT labeling was demonstrated in a wide dynamic range of 50 folds but without obvious matrix effect on quantification. Importantly, there was little quantification bias found among the individual IBT tags, indicating that the peptides labeled by different tags were quantitatively comparable. The IBT 10-plex reagents were applied for dynamically monitoring the quantitative responses of phosphoproteome ignited by EGF treatment in Hela cells. Total 5361 unique phosphopeptides were identified, which reached a similar conclusion as other reported. The IBT reagents were therefore experimentally proven as a new type of isobaric labeling peptides and useful for a large quantity peptides of quantitative proteomics.