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Title

High resolution reversed phase analysis of recombinant monoclonal antibodies by ultra-high pressure liquid chromatography column coupling

Authors
Dong, Michael W.
Zhang, Taylor
Published in Journal of Pharmaceutical and Biomedical Analysis. 2013, vol. 83, p. 273-278
Abstract Monoclonal antibodies (mAbs) represent one of the fastest growing areas of new drug development.However, their analytical characterization is complex and generally requires an array of orthogonal ana-lytical techniques. Reversed phase liquid chromatography is a valuable strategy due to its high resolvingpower and straightforward compatibility to mass spectrometry. The present study demonstrates thathigh peak capacity can be attained with intact mAb of ∼150 kDa, reduced mAb fragments of ∼25–50 kDaand also digested mAb generating numerous peptides of ∼0.5–3 kDa. Several long columns packed withfully porous wide-pore sub-2 µm particles were coupled in series to evaluate the effect of column lengthon peak capacity. By using three columns of 150 mm length, a mobile phase temperature of 80◦C and agradient time of around 20 min, peak capacities of 117 and 128 were obtained for a commercial intactmAb and its reduced mAb fragments, respectively. On the other hand, when peptide mapping was per-formed at 50◦C, with a gradient time of 270 min and a column length of 450 mm, a peak capacity of morethan 700 was achieved.
Keywords UHPLCHigh resolution separationsMonoclonal antibodiesmAbsPeak capacityHigh temperaturea
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Research group Sciences analytiques
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FEKETE, Szabolcs et al. High resolution reversed phase analysis of recombinant monoclonal antibodies by ultra-high pressure liquid chromatography column coupling. In: Journal of Pharmaceutical and Biomedical Analysis, 2013, vol. 83, p. 273-278. https://archive-ouverte.unige.ch/unige:28617

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Deposited on : 2013-06-17

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