In:
The Journal of Immunology, The American Association of Immunologists, Vol. 199, No. 12 ( 2017-12-15), p. 4180-4188
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 of Medium:
Online Resource
ISSN:
0022-1767
,
1550-6606
DOI:
10.4049/jimmunol.1701253
Language:
English
Publisher:
The American Association of Immunologists
Publication Date:
2017
detail.hit.zdb_id:
1475085-5
