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High pH reversed-phase fractionation profile of 100 µg HeLa cell lysate trypsic digest labeled with TMT reagent
Unique peptides (A) and protein groups (B) identified in each elution fraction compared to the total number of identifications from a combined search of all elution fractions and a single injection of unfractionated sample. Over 100% more unique peptides and over 50% more protein groups are identified in the sample upon high pH reversed-phase fractionation compared to analysis of an unfractionated sample. Excellent fractional resolution is attained (C), with only ~30% fractional overlap. The analysis, performed using triplicate sample sets, shows exceptional reproducibility, as indicated by the very narrow error bars.
