期刊
JOURNAL OF SEPARATION SCIENCE
卷 32, 期 8, 页码 1156-1164出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/jssc.200800615
关键词
High-pH RP chromatography; Low-pH ion-pair RP chromatography; Multidimensional separation; Repeatability; Shotgun proteome analysis; Strong cation-exchange chromatography; MS/MS
The repeatability of peptide identifications in shotgun proteome analyses employing strong cation-exchange x ion-pair RP HPLC hyphenated to ESI MS/MS was compared to an alternative scheme, comprising high-pH RP chromatography combined with low-pH ion-pair RP chromatography. Equivalent results were obtained with both methods in proteome analysis of Corynebacterium glutamicum. From a total number of 1350 to 1850 peptides identified in triplicate analyses of five consecutive fractions chosen from the first-dimension separation, 41-45% of the peptides were identified three times, whereas 16-22 and 37-39% of the peptides were identified only twice or once, respectively. A comparison of the repeatability of peptide identifications from complex samples upon 1- or 2-D chromatographic separation revealed that an additional separation dimension decreases the repeatability by approximately 25%.
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