10.1021/pr800318j.s002
Zhi-Bin Ning
Zhi-Bin
Ning
Qing-Run Li
Qing-Run
Li
Jie Dai
Jie
Dai
Rong-Xia Li
Rong-Xia
Li
Chia-Hui Shieh
Chia-Hui
Shieh
Rong Zeng
Rong
Zeng
Fractionation of Complex Protein Mixture by Virtual Three-Dimensional Liquid Chromatography Based on Combined pH and Salt Steps
American Chemical Society
2008
LTQ mass spectrometry
pH
protein mixtures
1933 protein groups
salt gradient elution
Salt StepsThe complexity
abundance proteins
cation exchange column
salt concentrations
phase column tandem
Complex Protein Mixture
Crude plasma
mass spectrometry identification
salt steps
cation exchange columns
desalted proteins
2008-10-03 00:00:00
Journal contribution
https://acs.figshare.com/articles/journal_contribution/Fractionation_of_Complex_Protein_Mixture_by_Virtual_Three_Dimensional_Liquid_Chromatography_Based_on_Combined_pH_and_Salt_Steps/2910163
The complexity and diversity of biological samples in proteomics require intensive fractionation ahead of mass spectrometry identification. This work developed a chromatographic method called virtual three-dimensional chromatography to fractionate complex protein mixtures. By alternate elution with different pHs and salt concentrations, we implemented pH and salt steps by turns on a single strong cation exchange column to fully exploit its chromatographic ability. Given standard proteins that were not resolved solely by pH or salt gradient elution could be successfully separated using this combined mode. With a reversed phase column tandem connected behind, we further fractionated as well as desalted proteins as the third dimension. This present strategy could readily be adapted with respect to special complexity of biological samples. Crude plasma without depleting high abundance proteins were fractionated by this three-dimensional mode and then analyzed by reversed phase liquid chromatography coupled with LTQ mass spectrometry. In total, 1933 protein groups with wide dynamic ranges were identified from a single experiment. Some characteristics that correlated to the behavior of proteins on strong cation exchange columns are also discussed.