Combination of COFRADIC and high temperature – extended column length conventional liquid chromatography: A very efficient way to tackle complex protein samples, such as serum

Autor: Lies Vanneste, Pat Sandra, Kris Gevaert, Filip D'hondt, Koen Sandra, Koen Kas, Grégoire Thomas, Christine Labeur, Katleen Verleysen, Joël Vandekerckhove
Rok vydání: 2007
Předmět:
Zdroj: Journal of Separation Science. 30:658-668
ISSN: 1615-9314
1615-9306
DOI: 10.1002/jssc.200600425
Popis: The previously reported COmbined FRActional DIagonal Chromatography (COFRA-DIC) methodology, in which a subset of peptides representative for their parent proteins are sorted, is particularly powerful for whole proteome analysis. This peptide-centric technology is built around diagonal chromatography, where peptide separations are crucial. This paper presents high efficiency peptide separations, in which four 250 x 2.1 mm, 5 microm Zorbax 300SB-C18 columns (total length 1 m) were coupled at operating temperatures of 60'C using a dedicated LC oven and conventional LC equipment. The high efficiency separations were combined with the COFRADIC procedure. This extremely powerful combination resulted, for the analysis of serum, in an increase in the uniquely identified peptide sequences by a factor of 2.6, compared to the COFRADIC procedure on a 25 cm column. This is a reflection of the increased peak capacity obtained on the 1 m column, which was calculated to be a factor 2.7 higher than on the 25 cm column. Besides more efficient sorting, less ion suppression was noticed.
Databáze: OpenAIRE
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