Format:
Online-Ressource
ISSN:
1522-2683
Content:
Abstract: A comprehensive two‐dimensional (2‐D) separation system, coupling capillary reverse‐phase liquid chromatography (cRPLC) to capillary isoelectric focusing (CIEF), is described for protein and peptide mapping. cRPLC, the first dimension, provided high‐resolution separations for salt‐free proteins. CIEF, the second dimension with an orthogonal mechanism to cRPLC afforded excellent resolution capability for proteins with efficient protein enrichment. Since all sample fractions in cRPLC effluents could be transferred to the CIEF dimensions, the combination of the two high‐efficiency separations resulted in maximal separation capabilities of each dimension. Separation effectiveness of this approach was demonstrated using complex protein/peptide samples, such as yeast cytosol and a BSA tryptic digest. A peak capacity of more than 10 000 had been achieved. A laser‐induced fluorescence (LIF) detector, developed for this system, allowed for high‐sensitive detection, with a fmol level of peptide detection for the BSA digest. FITC and BODIPY maleimide were used to tag the proteins, and the latter was found better both for separation and detection in our 2‐D system.
In:
volume:24
In:
number:18
In:
year:2003
In:
pages:3289-3295
In:
extent:7
In:
Electrophoresis, Weinheim : Wiley-Blackwell, 1980-, 24, Heft 18 (2003), 3289-3295 (gesamt 7), 1522-2683
Language:
English
DOI:
10.1002/elps.200305578
URN:
urn:nbn:de:101:1-2023100406513336826753
URL:
https://doi.org/10.1002/elps.200305578
URL:
https://nbn-resolving.org/urn:nbn:de:101:1-2023100406513336826753
URL:
https://d-nb.info/1304865401/34
URL:
https://doi.org/10.1002/elps.200305578
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