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Expert Rev Proteomics. 2012 Apr;9(2):129-34. doi: 10.1586/epr.12.15.

High-pH reversed-phase chromatography with fraction concatenation for 2D proteomic analysis.

Author information

1
Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA 99352, USA.

Abstract

Orthogonal high-resolution separations are critical for attaining improved analytical dynamic range and protein coverage in proteomic measurements. High-pH reversed-phase liquid chromatography (RPLC), followed by fraction concatenation, affords better peptide analysis than conventional strong cation-exchange chromatography applied for 2D proteomic analysis. For example, concatenated high-pH RPLC increased identification of peptides (by 1.8-fold) and proteins (by 1.6-fold) in shotgun proteomics analyses of a digested human protein sample. Additional advantages of high-pH RPLC with fraction concatenation include improved protein sequence coverage, simplified sample processing and reduced sample losses, making this an attractive alternative to strong cation-exchange chromatography in conjunction with second-dimension low-pH RPLC for 2D proteomics analyses.

PMID:
22462785
PMCID:
PMC3337716
DOI:
10.1586/epr.12.15
[Indexed for MEDLINE]
Free PMC Article

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