Pinder, CL;
Kratochvil, S;
Cizmeci, D;
Muir, L;
Guo, Y;
Shattock, RJ;
Mckay, PF;
(2017)
Isolation and Characterization of Antigen-Specific Plasmablasts Using a Novel Flow Cytometry–Based Ig Capture Assay.
Journal of Immunology
, 199
(12)
pp. 4180-4188.
10.4049/jimmunol.1701253.
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Muir_Isolation and Characterization of Antigen-Specific Plasmablasts Using a Novel Flow Cytometry-Based Ig Capture Assay_AAM.pdf - Accepted Version Access restricted to UCL open access staff Download (2MB) |
Abstract
We report the development of a novel flow cytometry–based Ig capture assay (ICA) for the identification and sorting of individual Ab-secreting cells based on their Ag reactivity. The ICA represents a fast and versatile tool for single-cell sorting of peripheral plasmablasts, streamlining subsequent Ab analysis, and cloning. We demonstrate the utility of the assay by isolating Ag-reactive plasmablasts from cryopreserved PBMC obtained from volunteers vaccinated with a recombinant HIV envelope protein. To show the specificity of the ICA, we produced Ag-specific Abs from these cells and subsequently verified their Ag reactivity via ELISA. Furthermore, we used the ICA to track Ag-specific plasmablast responses in HIV-vaccine recipients over a period of 42 d and performed a head-to-head comparison with a conventional B cell ELISpot. Results were highly comparable, highlighting that this assay is a viable alternative for monitoring Ag-specific plasmablast responses at early time points after infection or vaccination. The ICA provides important added benefits in that phenotypic information can be obtained from the identified Ag-specific cells that can then be captured for downstream applications such as B cell sequencing and/or Ab cloning. We envisage the ICA as being a useful tool in Ab repertoire analysis for future clinical trials.
| Type: | Article |
|---|---|
| Title: | Isolation and Characterization of Antigen-Specific Plasmablasts Using a Novel Flow Cytometry–Based Ig Capture Assay |
| DOI: | 10.4049/jimmunol.1701253 |
| Publisher version: | https://doi.org/10.4049/jimmunol.1701253 |
| Language: | English |
| Additional information: | This version is the author accepted manuscript. For information on re-use, please refer to the publisher’s terms and conditions. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Infection and Immunity |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10071899 |
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