Label-Free Glycoprofiling with Multiplex Surface Plasmon Resonance: A Tool To Quantify Sialylation of Erythropoietin

K.P.M. Geuijen, L.A. Halims, H. Schellekens, R.B. Schasfoort, R.H. Wijffels, M.H.M. Eppink

Research output: Contribution to journalArticleAcademicpeer-review

16 Citations (Scopus)

Abstract

Protein glycosylation is among the most common and well-defined post-translational modifications due to its vital role in protein function. Monitoring variation in glycosylation is necessary for producing more effective therapeutic proteins. Glycans attached to glycoproteins interact highly specific with lectins, natural carbohydrate-binding proteins, which property is used in the current label-free methodology. We have established a lectin microarray for label-free detection of lectin-carbohydrate interactions allowing us to study protein glycosylation directly on unmodified glycoproteins. The method enables simultaneous measurement of up to 96 lectin-carbohydrate interactions on a multiplex surface plasmon resonance imaging platform within 20 min. Specificity determination of lectins succeeded by analysis of neoglycoproteins and enzymatically remodeled glycoproteins to verify carbohydrate binding. We demonstrated the possibilities for glycosylation fingerprinting by comparing different Erythropoietin sources without the need for any sample pretreatment and we were able to accurately quantify relative sialylation levels of Erythropoietin.
Original languageEnglish
Pages (from-to)8115-8122
JournalAnalytical Chemistry
Volume87
Issue number16
DOIs
Publication statusPublished - 2015

Keywords

  • liquid chromatography/mass spectrometry
  • recombinant-human-erythropoietin
  • lectin microarray
  • biological function
  • real-time
  • cho-cells
  • glycosylation
  • carbohydrate
  • glycoproteins
  • hemoglobin

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