De Novo MS/MS Sequencing of Native Human Antibodies
peer-reviewed · Journal of Proteome Research · 2017
peer-reviewed · Journal of Proteome Research · 2017. Adrian Guthals et al. One direct route for the discovery of therapeutic human monoclonal antibodies (mAbs) involves the isolation…
| Date | 2017-01-06 |
| Type | peer-reviewed |
| Venue | Journal of Proteome Research |
| Publisher | American Chemical Society (ACS) |
| Contribution | downstream-application |
| DOI | 10.1021/acs.jproteome.6b00608 |
| Citations (OpenAlex) | 60 |
| Venue 2-year citedness | 3.83 |
Abstract
One direct route for the discovery of therapeutic human monoclonal antibodies (mAbs) involves the isolation of peripheral B cells from survivors/sero-positive individuals after exposure to an infectious reagent or disease etiology, followed by single-cell sequencing or hybridoma generation. Peripheral B cells, however, are not always easy to obtain and represent only a small percentage of the total B-cell population across all bodily tissues. Although it has been demonstrated that tandem mass spectrometry (MS/MS) techniques can interrogate the full polyclonal antibody (pAb) response to an antigen in vivo, all current approaches identify MS/MS spectra against databases derived from genetic sequencing of B cells from the same patient. In this proof-of-concept study, we demonstrate the feasibility of a novel MS/MS antibody discovery approach in which only serum antibodies are required without the need for sequencing of genetic material. Peripheral pAbs from a cytomegalovirus-exposed individual were purified by glycoprotein B antigen affinity and de novo sequenced from MS/MS data. Purely MS-derived mAbs were then manufactured in mammalian cells to validate potency via antigen-binding ELISA. Interestingly, we found that these mAbs accounted for 1 to 2% of total donor IgG but were not detected in parallel sequencing of memory B cells from the same patient.
Methods and tools
- Native antibody de novo MS/MS: De novo MS/MS sequencing workflow for native human antibodies.
Data deposited
- De Novo MS/MS Sequencing of Native Human Antibodies (as deposited) · MSV000080039
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