Comparison of displacement versus gradient mode for separation of a complex protein mixture by anion-exchange chromatography

Standard

Comparison of displacement versus gradient mode for separation of a complex protein mixture by anion-exchange chromatography. / Ahrends, Robert; Lichtner, Björn; Buck, Friedrich; Hildebrand, Diana; Kotasinska, Marta; Kohlbacher, Oliver; Kwiatkowski, Marcel; Wagner, Moritz; Trusch, Maria; Schlüter, Hartmut.

In: J CHROMATOGR B, Vol. 901, 15.07.2012, p. 34-40.

Research output: SCORING: Contribution to journalSCORING: Journal articleResearchpeer-review

Harvard

Ahrends, R, Lichtner, B, Buck, F, Hildebrand, D, Kotasinska, M, Kohlbacher, O, Kwiatkowski, M, Wagner, M, Trusch, M & Schlüter, H 2012, 'Comparison of displacement versus gradient mode for separation of a complex protein mixture by anion-exchange chromatography', J CHROMATOGR B, vol. 901, pp. 34-40. https://doi.org/10.1016/j.jchromb.2012.05.037

APA

Ahrends, R., Lichtner, B., Buck, F., Hildebrand, D., Kotasinska, M., Kohlbacher, O., Kwiatkowski, M., Wagner, M., Trusch, M., & Schlüter, H. (2012). Comparison of displacement versus gradient mode for separation of a complex protein mixture by anion-exchange chromatography. J CHROMATOGR B, 901, 34-40. https://doi.org/10.1016/j.jchromb.2012.05.037

Vancouver

Bibtex

@article{bac8f1bebc3a4e02a31c6f021e2f3570,
title = "Comparison of displacement versus gradient mode for separation of a complex protein mixture by anion-exchange chromatography",
abstract = "Liquid chromatography is often the method of choice for the analysis of proteins in their native state. Nevertheless compared to two-dimensional electrophoresis, the resolution of common chromatographic techniques is low. Liquid chromatography in the displacement mode has previously been shown to offer higher resolution and to elute proteins in the high concentrations. In this study we compared to what extend displacement mode was a suitable alternative to gradient mode for the separation of a complex protein mixture using anion-exchange displacement chromatography and if it is therefore helpful for proteomic investigations. Hence we analyzed the qualitative protein composition of each fraction by tryptic digestion of the proteins, analysis of the tryptic peptides by liquid chromatography coupled to mass spectrometry followed by data base analysis and by measuring the elution profiles of 22 selected proteins with selected reaction monitoring mass spectrometry. In the fractions of displacement mode a significantly higher number of identified proteins (51 versus 16) was yielded in comparison to gradient mode. The resolution of displacement chromatography was slightly lower than of gradient chromatography for many but not for all proteins. The selectivities of displacement mode and gradient mode are very different. In conclusion displacement chromatography is a well suited alternative for top-down proteomic approaches which start with separating intact proteins first prior to mass spectrometric analysis of intact or digested proteins. The significant orthogonality of both modes may be used in the future for combining them in multidimensional fractionation procedures.",
keywords = "Humans, Mass Spectrometry/*methods, Anions/chemistry, Blood Proteins/chemistry/*isolation & purification, Chromatography, Ion Exchange/*methods, Electric Conductivity, Spectrophotometry, Ultraviolet, Humans, Mass Spectrometry/*methods, Anions/chemistry, Blood Proteins/chemistry/*isolation & purification, Chromatography, Ion Exchange/*methods, Electric Conductivity, Spectrophotometry, Ultraviolet",
author = "Robert Ahrends and Bj{\"o}rn Lichtner and Friedrich Buck and Diana Hildebrand and Marta Kotasinska and Oliver Kohlbacher and Marcel Kwiatkowski and Moritz Wagner and Maria Trusch and Hartmut Schl{\"u}ter",
note = "Copyright {\textcopyright} 2012 Elsevier B.V. All rights reserved.",
year = "2012",
month = jul,
day = "15",
doi = "10.1016/j.jchromb.2012.05.037",
language = "English",
volume = "901",
pages = "34--40",
journal = "J CHROMATOGR B",
issn = "1570-0232",
publisher = "Elsevier",

}

RIS

TY - JOUR

T1 - Comparison of displacement versus gradient mode for separation of a complex protein mixture by anion-exchange chromatography

AU - Ahrends, Robert

AU - Lichtner, Björn

AU - Buck, Friedrich

AU - Hildebrand, Diana

AU - Kotasinska, Marta

AU - Kohlbacher, Oliver

AU - Kwiatkowski, Marcel

AU - Wagner, Moritz

AU - Trusch, Maria

AU - Schlüter, Hartmut

N1 - Copyright © 2012 Elsevier B.V. All rights reserved.

PY - 2012/7/15

Y1 - 2012/7/15

N2 - Liquid chromatography is often the method of choice for the analysis of proteins in their native state. Nevertheless compared to two-dimensional electrophoresis, the resolution of common chromatographic techniques is low. Liquid chromatography in the displacement mode has previously been shown to offer higher resolution and to elute proteins in the high concentrations. In this study we compared to what extend displacement mode was a suitable alternative to gradient mode for the separation of a complex protein mixture using anion-exchange displacement chromatography and if it is therefore helpful for proteomic investigations. Hence we analyzed the qualitative protein composition of each fraction by tryptic digestion of the proteins, analysis of the tryptic peptides by liquid chromatography coupled to mass spectrometry followed by data base analysis and by measuring the elution profiles of 22 selected proteins with selected reaction monitoring mass spectrometry. In the fractions of displacement mode a significantly higher number of identified proteins (51 versus 16) was yielded in comparison to gradient mode. The resolution of displacement chromatography was slightly lower than of gradient chromatography for many but not for all proteins. The selectivities of displacement mode and gradient mode are very different. In conclusion displacement chromatography is a well suited alternative for top-down proteomic approaches which start with separating intact proteins first prior to mass spectrometric analysis of intact or digested proteins. The significant orthogonality of both modes may be used in the future for combining them in multidimensional fractionation procedures.

AB - Liquid chromatography is often the method of choice for the analysis of proteins in their native state. Nevertheless compared to two-dimensional electrophoresis, the resolution of common chromatographic techniques is low. Liquid chromatography in the displacement mode has previously been shown to offer higher resolution and to elute proteins in the high concentrations. In this study we compared to what extend displacement mode was a suitable alternative to gradient mode for the separation of a complex protein mixture using anion-exchange displacement chromatography and if it is therefore helpful for proteomic investigations. Hence we analyzed the qualitative protein composition of each fraction by tryptic digestion of the proteins, analysis of the tryptic peptides by liquid chromatography coupled to mass spectrometry followed by data base analysis and by measuring the elution profiles of 22 selected proteins with selected reaction monitoring mass spectrometry. In the fractions of displacement mode a significantly higher number of identified proteins (51 versus 16) was yielded in comparison to gradient mode. The resolution of displacement chromatography was slightly lower than of gradient chromatography for many but not for all proteins. The selectivities of displacement mode and gradient mode are very different. In conclusion displacement chromatography is a well suited alternative for top-down proteomic approaches which start with separating intact proteins first prior to mass spectrometric analysis of intact or digested proteins. The significant orthogonality of both modes may be used in the future for combining them in multidimensional fractionation procedures.

KW - Humans

KW - Mass Spectrometry/methods

KW - Anions/chemistry

KW - Blood Proteins/chemistry/isolation & purification

KW - Chromatography, Ion Exchange/methods

KW - Electric Conductivity

KW - Spectrophotometry, Ultraviolet

KW - Humans

KW - Mass Spectrometry/methods

KW - Anions/chemistry

KW - Blood Proteins/chemistry/isolation & purification

KW - Chromatography, Ion Exchange/methods

KW - Electric Conductivity

KW - Spectrophotometry, Ultraviolet

U2 - 10.1016/j.jchromb.2012.05.037

DO - 10.1016/j.jchromb.2012.05.037

M3 - SCORING: Journal article

C2 - 22727752

VL - 901

SP - 34

EP - 40

JO - J CHROMATOGR B

JF - J CHROMATOGR B

SN - 1570-0232

ER -