The Use of Ammonium Formate as a Mobile-Phase Modifier for LC-MS/MS Analysis of Tryptic Digests
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TLDR
The use of FA/AF improved online RP-LC separations and led to significant increases in peptide identifications with improved protein sequence coverage.Abstract:
A major challenge facing current mass spectrometry (MS)-based proteomics research is the large concentration range displayed in biological systems, which far exceeds the dynamic range of commonly available mass spectrometers. One approach to overcome this limitation is to improve online reversed-phase liquid chromatography (RP-LC) separation methodologies. LC mobile-phase modifiers are used to improve peak shape and increase sample load tolerance. Trifluoroacetic acid (TFA) is a commonly used mobile-phase modifier, as it produces peptide separations that are far superior to other additives. However, TFA leads to signal suppression when incorporated with electrospray ionization (ESI), and thus, other modifiers, such as formic acid (FA), are used for LC-MS applications. FA exhibits significantly less signal suppression, but is not as effective of a modifier as TFA. An alternative mobile-phase modifier is the combination of FA and ammonium formate (AF), which has been shown to improve peptide separations. The ESI-MS compatibility of this modifier has not been investigated, particularly for proteomic applications. This work compares the separation metrics of mobile phases modified with FA and FA/AF and explores the use of FA/AF for the LC-MS analysis of tryptic digests. Standard tryptic-digest peptides were used for comparative analysis of peak capacity and sample load tolerance. The compatibility of FA/AF in proteomic applications was examined with the analysis of soluble proteins from canine prostate carcinoma tissue. Overall, the use of FA/AF improved online RP-LC separations and led to significant increases in peptide identifications with improved protein sequence coverage.read more
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References
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Journal ArticleDOI
The Human Plasma Proteome History, Character, and Diagnostic Prospects
TL;DR: This work speculates on the reasons behind this large discrepancy between the expectations arising from proteomics and the realities of clinical diagnostics and suggests approaches by which protein-disease associations may be more effectively translated into diagnostic tools in the future.
Journal ArticleDOI
Mass spectrometry and protein analysis.
TL;DR: Recent advances in mass spectrometry instrumentation are reviewed in the context of current and emerging research strategies in protein science.
Journal ArticleDOI
Enhanced sensitivity for peptide mapping with electrospray liquid chromatography-mass spectrometry in the presence of signal suppression due to trifluoroacetic acid-containing mobile phases
TL;DR: A method is described for improving the sensitivity of peptide mapping with electrospray liquid chromatography--mass spectrometry using trifluoroacetic acid (TFA) containing HPLC mobile phases and the enhanced sensitivity allowed location of a modified residue by comparison endoproteinase Lys C digest of native and oxidized forms of the protein without extensive sample preparation or concentration.
Journal ArticleDOI
Fast analysis in liquid chromatography using small particle size and high pressure
TL;DR: Advantages and problems encountered with high pressure in terms of frictional heating and solvent compressibility will be discussed even if systems working at a maximum pressure of 1000 bar are not influenced by these parameters and give reliable and reproducible results.
Journal ArticleDOI
The challenges of the analysis of basic compounds by high performance liquid chromatography: some possible approaches for improved separations.
TL;DR: This review considers some of the difficulties encountered with the analysis of ionised bases using reversed-phase chromatography, such as detrimental interaction with column silanol groups, and overloading which both lead to poor peak shapes.