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J Chromatogr A. 2017 Nov 10;1523:283-292. doi: 10.1016/j.chroma.2017.06.052. Epub 2017 Jun 19.

The versatility of heart-cutting and comprehensive two-dimensional liquid chromatography in monoclonal antibody clone selection.

Author information

1
Research Institute for Chromatography (RIC), President Kennedypark 26, B-8500 Kortrijk, Belgium; anaRIC biologics, Noorwegenstraat 4, B-9940 Evergem, Ghent, Belgium. Electronic address: koen.sandra@richrom.com.
2
Research Institute for Chromatography (RIC), President Kennedypark 26, B-8500 Kortrijk, Belgium.
3
Research Institute for Chromatography (RIC), President Kennedypark 26, B-8500 Kortrijk, Belgium; anaRIC biologics, Noorwegenstraat 4, B-9940 Evergem, Ghent, Belgium.

Abstract

In recent years, two-dimensional liquid chromatography (2D-LC) has seen an enormous evolution and one of the fields where it is being widely adopted is in the analysis of therapeutic monoclonal antibodies (mAbs). We here further add to the many flavours of this powerful technology. Workflows based on heart-cutting (LC-LC) and comprehensive (LC×LC) 2D-LC are described that allow to guide the clone selection process in mAb and biosimilar development. Combining Protein A affinity chromatography in the first dimension with size exclusion (SEC), cation exchange (CEX) or reversed-phase liquid chromatography-mass spectrometry (RPLC-MS) in the second dimension simultaneously allows to assess mAb titer and critical structural aspects such as aggregation, fragmentation, charge heterogeneity, molecular weight (MW), amino acid sequence and glycosylation. Complementing the LC-LC measurements at intact protein level with LC×LC based peptide mapping provides the necessary information to make clear decisions on which clones to take further into development.

KEYWORDS:

Biosimilar; Clone selection; Monoclonal antibody; Protein A; Two-dimensional liquid chromatography

PMID:
28668371
DOI:
10.1016/j.chroma.2017.06.052
[Indexed for MEDLINE]

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