Characterization of human plasma glycoproteins separated by two-dimensional gel electrophoresis

Nicolle H. Packer, Marc R. Wilkins, Olivier Golaz, Margaret A. Lawson, Andrew A. Gooley, Denis F. Hochstrasser, John W. Redmond, Keith L. Williams*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

37 Citations (Scopus)


Purification of protein isoforms for the characterization of post-translational modifications, such as glycosylation, can be laborious and demanding. We report a means of determining monosaccharide composition and the identity of glycoproteins from a single spot on a two-dimensional (2-D) gel. The sensitivity of the method depends on the degree of glycosylation of the protein. We show that bovine fetuin can be analyzed and identified at the level of 100 pmol. 2-D reference maps enable quick identification of glycoprotein isoforms, and the nature of glycosylation differences. Human sera glycoforms were isolated by micropreparative 2-D PAGE using a narrow-range immobilized pH gradient. Single spots excised from one polyvinylidene difluoride blot of a 2-D gel were used sequentially for sialic acid analysis, neutral and amino sugar analysis, and finally amino acid analysis. The glycosylation variations in isoforms of human fetuin and α-1-antitrypsin were determined. The amino acid composition, in conjunction with protein pI and MW, successfully identified the glycoproteins.

Original languageEnglish
Pages (from-to)66-70
Number of pages5
Issue number1
Publication statusPublished - 1996


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