Related Experiment Video
Updated: Jul 7, 2026

Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
Published on: November 15, 2017
Evaluation of comprehensive multidimensional separations using reversed-phase, reversed-phase liquid
Tatsuji Nakamura1, Junro Kuromitsu, Yoshiya Oda
1Eisai Company Ltd., Laboratory of Core Technology, Ibaraki, Japan.
Abstract:
Two-dimensional liquid-chromatographic (LC) separation followed by mass spectrometric (MS) analysis was examined for the identification of peptides in complex mixtures as an alternative to widely used two-dimensional gel electrophoresis followed by MS analysis for use in proteomics. The present method involves the off-line coupling of a narrow-bore, polymer-based, reversed-phase column using an acetonitrile gradient in an alkaline mobile phase in the first dimension with octadecylsilanized silica (ODS)-based nano-LC/MS in the second dimension. After the first separation, successive fractions were acidified and dried off-line, then loaded on the second dimension column. Both columns separate peptides according to hydrophobicity under different pH conditions, but more peptides were identified than with the conventional technique for shotgun proteomics, that is, the combination of a strong cation exchange column with an ODS column, and the system was robust because no salts were included in the mobile phases. The suitability of the method for proteomics measurements was evaluated.
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...
Tandem Mass Spectrometry
Mass Spectrometry: Complex Analysis
GC–MS is a powerful hyphenated method commonly used in forensics and environmental...

