Separation techniques hyphenated to electrospray-tandem mass spectrometry in proteomics: capillary electrophoresis versus nanoliquid chromatography.
Liquid chromatography (LC) nanoelectrospray-tandem mass spectrometry (MS/MS) is a key technology for the study of proteomics, with the main benefit to the characterization of sensitive peptides from complex mixtures. Capillary electrophoresis coupled to mass spectrometry (MS) has been taken into consideration sporadically due to the highly efficient separation and ability to handle low sample amount, yet classified as being less sensitive with respect to analyte concentration. The limitation in capillary zone electrophoresis (CZE) injection volumes can be overcome by on-line solid-phase extraction (SPE). Such an on-line SPE-CZE system was explored in combination with an ion trap (IT) mass spectrometer. Thus, it was possible to inject more than 100 microL sample solution on to the CZE capillary. Concentration limits of detection as low as 100 amol/microL were demonstrated for a peptide standard. This SPE-CZE-microelectrospray ionization (ESI)-MS/MS setup was compared directly to nanoLC/nanoESI using the same sample of a tryptic digest of bovine serum albumin (BSA) as a reference standard. Measurements were made on one IT mass spectrometer with identical acquisition parameters. Both chromatography systems enabled the separation and detection of low levels of peptides from a mixture of moderate complexity, with most peptides identified using both techniques; however, specific differences were obvious. The nanoLC-MS is about five times more sensitive than the CZE-MS, yet the difference was less pronounced than expected. The CZE-MS technique showed reduced loss of peptides, especially for larger peptides (missed cleavages) and is about four times faster than the nanoLC-MS approach.
Pelzing
Matthias
M
Bruker Daltonik GmbH, Leipzig, Germany. mp@bdal.de
Neusüss
Christian
C
D000595:Amino Acid Sequence; D002853:Chromatography, Liquid / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D008969:Molecular Sequence Data; D036103:Nanotechnology; D010455:Peptides / Q000302:isolation & purification; D040901:Proteomics / Q000295:instrumentation*; D012680:Sensitivity and Specificity; D020539:Sequence Analysis, Protein; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D010455:Peptides