Complementary Methods for de Novo Monoclonal Antibody Sequencing to Achieve Complete Sequence Coverage

Mass spectrometry is a powerful tool for de novo sequencing of novel proteins. Recent efforts in this area have mainly focused on liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Here, we present an alternative method, capillary electrophoresis tandem mass spectrometry (CE-MS/...

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Bibliographic Details
Published in:Journal of proteome research 2020-07, Vol.19 (7), p.2700-2707
Main Authors: Cheng, Jianhui, Wang, Lingyu, Rive, Craig M, Holt, Robert A, Morin, Gregg B, Chen, David D. Y
Format: Article
Language:English
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Summary:Mass spectrometry is a powerful tool for de novo sequencing of novel proteins. Recent efforts in this area have mainly focused on liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Here, we present an alternative method, capillary electrophoresis tandem mass spectrometry (CE-MS/MS), for sequencing novel monoclonal antibodies. Using less than 200 ng in total of tryptic digest sample in a triplicated measurement, CE-MS/MS with pH-mediated focusing successfully sequenced mAb infliximab with 100% sequence coverage and 100% accuracy for the light chain and 96% coverage and 93% accuracy for the heavy chain. It was also demonstrated that CE-MS/MS gives comparable results, and in some cases, even better results, as compared to LC-MS/MS when used as a standalone technique. A combined workflow using both CE-MS/MS and LC-MS/MS was also used to sequence a novel antibody, anti-CD-176, resulting in the first proposed sequence for this mAb.
ISSN:1535-3893
1535-3907
DOI:10.1021/acs.jproteome.0c00223