Glycoform analysis of Alpha1-Acid glycoprotein by capillary electrophoresis using electrophoretic injection

Chenhua Zhang, William Clarke, David S. Hage

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Human alpha1-acid glycoprotein (AGP) is an acute phase glycoprotein that has a heterogeneous glycosylation pattern. This pattern can change in certain diseases, which has resulted in interest in using AGP glycoforms as potential biomarkers for these diseases. This report describes a method that uses capillary electrophoresis to characterize and analyze AGP glycoforms both in purified samples of AGP and in human serum. This method uses static and dynamic coatings of poly(ethylene oxide) that are applied to a silica capillary for separation of AGP glycoforms in the reversed-polarity mode of CE and in the presence of negligible electroosmotic flow. Electrophoretic injection is performed onto such capillaries by using a stationary stacking interface between the sample and running buffer. In addition, acidic precipitation and desalting are used to allow for the isolation and the analysis of AGP from only 65 μL of serum. Up to eleven AGP glycoform bands can be reproducibly separated by this method, with the difference in migration time between neighboring bands being 12- to almost 60-fold larger than the standard deviation for the migration time of any given band. A limit of detection down to about 2 nM per glycoform band can be obtained by this method for AGP in serum based on absorbance detection and without the need for further sample modification or labeling.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages41-56
Number of pages16
DOIs
StatePublished - Jan 1 2019

Publication series

NameMethods in Molecular Biology
Volume1972
ISSN (Print)1064-3745

Fingerprint

Capillary Electrophoresis
Glycoproteins
Injections
Acids
Serum
Electroosmosis
Ethylene Oxide
Acute-Phase Proteins
Glycosylation
Silicon Dioxide
Limit of Detection
Buffers
Biomarkers

Keywords

  • Alpha-acid glycoprotein
  • Capillary electrophoresis
  • Capillary modification
  • Electrophoretic injection
  • Glycoforms
  • Sample stacking
  • Serum

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Zhang, C., Clarke, W., & Hage, D. S. (2019). Glycoform analysis of Alpha1-Acid glycoprotein by capillary electrophoresis using electrophoretic injection. In Methods in Molecular Biology (pp. 41-56). (Methods in Molecular Biology; Vol. 1972). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-9213-3_4

Glycoform analysis of Alpha1-Acid glycoprotein by capillary electrophoresis using electrophoretic injection. / Zhang, Chenhua; Clarke, William; Hage, David S.

Methods in Molecular Biology. Humana Press Inc., 2019. p. 41-56 (Methods in Molecular Biology; Vol. 1972).

Research output: Chapter in Book/Report/Conference proceedingChapter

Zhang, C, Clarke, W & Hage, DS 2019, Glycoform analysis of Alpha1-Acid glycoprotein by capillary electrophoresis using electrophoretic injection. in Methods in Molecular Biology. Methods in Molecular Biology, vol. 1972, Humana Press Inc., pp. 41-56. https://doi.org/10.1007/978-1-4939-9213-3_4
Zhang C, Clarke W, Hage DS. Glycoform analysis of Alpha1-Acid glycoprotein by capillary electrophoresis using electrophoretic injection. In Methods in Molecular Biology. Humana Press Inc. 2019. p. 41-56. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-4939-9213-3_4
Zhang, Chenhua ; Clarke, William ; Hage, David S. / Glycoform analysis of Alpha1-Acid glycoprotein by capillary electrophoresis using electrophoretic injection. Methods in Molecular Biology. Humana Press Inc., 2019. pp. 41-56 (Methods in Molecular Biology).
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