High-Recovery Desalting Tip Columns for a Wide Variety of Peptides in Mass Spectrometry-based Proteomics

Abstract

ABSTRACTIn mass spectrometry-based proteomics, loss-minimized peptide purification techniques play a key role in improving sensitivity and coverage. We have developed a desalting tip column packed with thermoplastic polymer-coated chromatographic particles, named ChocoTip, to achieve high recoveries in peptide purification by pipette-tip-based LC with centrifugation (tipLC). ChocoTip identified more than twice as many peptides from 20 ng of tryptic peptides from Hela cell lysate compared to a typical StageTip packed with chromatographic particles entangled in a Teflon mesh in tipLC. The high recovery of ChocoTip in tipLC was maintained for peptides with a wide variety of physical properties over the entire retention time range of the LC/MS/MS analysis, and was especially noteworthy for peptides with long retention times. These excellent properties are attributable to the unique morphology of ChocoTip, in which the thermoplastic polymer covers the pores, thereby inhibiting irreversible adsorption of peptides into mesopores of the chromatographic particles.ChocoTip is expected to find applications, especially in clinical proteomics and single-cell proteomics, where sample amounts are limited.