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Available under license: CC BY: Creative Commons Attribution 4.0 International License
Final published version
Research output: Contribution to Journal/Magazine › Journal article › peer-review
Research output: Contribution to Journal/Magazine › Journal article › peer-review
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TY - JOUR
T1 - Detection of protein glycosylation using tip enhanced Raman scattering
AU - Cowcher, David
AU - Deckert-Gaudig, Tanja
AU - Brewster, Victoria
AU - Ashton, Lorna
AU - Deckert, Volker
AU - Goodacre, Royston
N1 - ACS AuthorChoice - This is an open access article published under a Creative Commons Attribution (CC-BY) License, which permits unrestricted use, distribution and reproduction in any medium, provided the author and source are cited.
PY - 2016/2/16
Y1 - 2016/2/16
N2 - The correct glycosylation of biopharmaceutical glycoproteins and their formulations is essential for them to have the desired therapeutic effect on the patient. It has recently been shown that Raman spectroscopy can be used to quantify the proportion of glycosylated protein from mixtures of native and glycosylated forms of bovine pancreatic ribonuclease (RNase). Here we show the first steps towards not only the detection of glycosylation status, but the characterisation of glycans themselves from just a few protein molecules at a time using tip-enhanced Raman scattering (TERS). Whilst this technique generates complex data that are very dependent on the protein orientation, with the careful development of combined data preprocessing, univariate and multivariate analysis techniques, we have shown that we can distinguish between the native and glycosylated forms of RNase. Many glycoproteins contain populations of subtly different glycoforms, therefore with stricter orientation control, we believe this has the potential to lead to further glycan characterisation using TERS, which would have use in biopharmaceutical synthesis and formulation research.
AB - The correct glycosylation of biopharmaceutical glycoproteins and their formulations is essential for them to have the desired therapeutic effect on the patient. It has recently been shown that Raman spectroscopy can be used to quantify the proportion of glycosylated protein from mixtures of native and glycosylated forms of bovine pancreatic ribonuclease (RNase). Here we show the first steps towards not only the detection of glycosylation status, but the characterisation of glycans themselves from just a few protein molecules at a time using tip-enhanced Raman scattering (TERS). Whilst this technique generates complex data that are very dependent on the protein orientation, with the careful development of combined data preprocessing, univariate and multivariate analysis techniques, we have shown that we can distinguish between the native and glycosylated forms of RNase. Many glycoproteins contain populations of subtly different glycoforms, therefore with stricter orientation control, we believe this has the potential to lead to further glycan characterisation using TERS, which would have use in biopharmaceutical synthesis and formulation research.
U2 - 10.1021/acs.analchem.5b03535
DO - 10.1021/acs.analchem.5b03535
M3 - Journal article
VL - 88
SP - 2105
EP - 2112
JO - Analytical Chemistry
JF - Analytical Chemistry
SN - 0003-2700
IS - 4
ER -