6533b873fe1ef96bd12d572b
RESEARCH PRODUCT
Enhancing Sensitivity of Microflow-Based Bottom-Up Proteomics through Postcolumn Solvent Addition.
Sven SchumannMarkus WanningerStefan TenzerMateusz Krzysztof ŁąCkiUte Distlersubject
ProteomicsElectrospraySpectrometry Mass Electrospray IonizationChromatographyChemistryDimethyl sulfoxideElectrospray ionization010401 analytical chemistry010402 general chemistryMass spectrometry01 natural sciences0104 chemical sciencesAnalytical ChemistrySolventchemistry.chemical_compoundSolventsHumansNanotechnologyDimethyl SulfoxideBottom-up proteomicsMethanolAcetonitrilePeptidesChromatography High Pressure LiquidHeLa Cellsdescription
The introduction of more sensitive mass spectrometers allows researchers to adapt front-end liquid chromatography (LC) to individual needs for the analysis of complex proteomes. Where absolute sensitivity is not paramount, it is advantageous to switch from a highly sensitive nanoflow-LC setup, the de facto standard platform in mass-spectrometry (MS)-based discovery proteomics, to a more robust, high-throughput-compatible microflow or conventional-flow setup. To enhance the microflow-LC-MS electrospray process of complex proteomic samples, we tested the effects of different solvents, including 2-propanol, methanol, and acetonitrile, pure or as mixture with dimethyl sulfoxide, which were added postcolumn to the eluting sample. Postcolumn addition of organic solvents strongly enhanced the electrospray efficiency in microflow-LC-MS experiments and improved the sensitivity across the entire gradient and for early eluting peptides by up to 10-fold. Postcolumn solvent addition did not negatively affect chromatographic performance and resulted in an overall 28-36% increase in identifications at both the protein and peptide levels. The presented microflow-LC-MS workflow, including postcolumn solvent addition, can be easily adopted on any LC-MS/MS platform.
| year | journal | country | edition | language |
|---|---|---|---|---|
| 2019-05-22 | Analytical chemistry |