Quantitative characterization of intact sialylated O-glycans with MALDI-MS for protein biotherapeutics
- 56 Downloads
For validating O-glycosylation of protein biotherapeutics, we presented a quantitative O-glycomics method which is based on the neutralization of sialic acids, the specific release of O-glycans, and the introduction of permanent positive charge followed by quantitative MALDI-MS analysis. This method shows excellent technical reproducibility, linearity and sensitivity. In addition, it enables the quantification of intact O-glycans with minimal degradation or loss of sialic acids on these glycans compared to a conventional HPLC-based method. We then applied this method to quantitatively characterize O-glycans present on Etanercept. The analysis showed the relative abundances of mono- and di-sialylated core 1 O-glycans - were 79.3±0.8% and 17.3±1.4%, respectively. This glycomics technology could allow for the reliable quantitative analysis of intact O-glycans from glycoproteins and may contribute to validation of O-glycosylation protein biotherapeutics in the pharmaceutical industry.
KeywordsChemical Derivatization MALDI-MS O-glycan Protein Biotherapeutics Quantitative Analysis
Unable to display preview. Download preview PDF.
- 1.S. Elliott, T. Lorenzini, S. Asher, K. Aoki, D. Brankow, L. Buck, L. Busse, D. Chang, J. Fuller, J. Grant, N. Hernday, M. Hokum, S. Hu, A. Knudten, N. Levin, R. Komorowski, F. Martin, R. Navarro, T. Osslund, G. Rogers, N. Rogers, G. Trail and J. Egrie, Nat. Biotechnol., 21, 414 (2003).CrossRefPubMedGoogle Scholar