Related Experiment Video
Updated: Aug 21, 2026

Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
Published on: November 15, 2017
Physicochemical properties determining the detection probability of tryptic peptides in Fourier transform mass
Michael L Nielsen1, Mikhail M Savitski, Frank Kjeldsen
1Laboratory for Biological and Medical Mass Spectrometry, Uppsala University, Sweden. Michael-Lund-Nielsen@bmms.uu.se
Abstract:
Sequence verification and mapping of posttranslational modifications require nearly 100% sequence coverage in the "bottom-up" protein analysis. Even in favorable cases, routine liquid chromatography-mass spectrometry detects from protein digests peptides covering 50-90% of the sequence. Here we investigated the reasons for limited peptide detection, considering various physicochemical aspects of peptide behavior in liquid chromatography-Fourier transform mass spectrometry (LC-FTMS). No overall correlation was found between the detection probability and peptide mass. In agreement with literature data, the signal increased with peptide hydrophobicity. Surprisingly, the pI values exhibited an opposite trend, with more acidic tryptic peptides detected with higher probability. A mixture of synthesized peptides of similar masses confirmed the hydrophobicity dependence but showed strong positive correlation between pI and signal response. An explanation of this paradoxal behavior was found through the observation that more acidic tryptic peptide lengths tend to be longer. Longer peptides tend to acquire higher average charge state in positive mode electrospray ionization than more basic but shorter counterparts. The induced-current detection in FTMS favors ions in higher charge states, thus providing the observed pI-FTMS signal anticorrelation.
Related Concept Videos
Peptide Identification Using Tandem Mass Spectrometry
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...
MALDI-TOF Mass Spectrometry
Mass Spectrum: Interpretation
Mass Spectrometry: Complex Analysis
GC–MS is a powerful hyphenated method commonly used in forensics and environmental...
High-Performance Liquid Chromatography: Types of Detectors
Mass Spectrometry: Carboxylic Acid, Ester, and Amide Fragmentation
For example, the fragmentation of...
