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J Vis Exp. 2017 Feb 16;(120). doi: 10.3791/55382.

Detection and Enrichment of Rare Antigen-specific B Cells for Analysis of Phenotype and Function.

Author information

1
Department of Immunology and Microbiology, University of Colorado School of Medicine; Department of Microbiology, Immunology, and Pathology, Colorado State University.
2
Department of Immunology and Microbiology, University of Colorado School of Medicine.
3
Barbara Davis Center for Childhood Diabetes, University of Colorado School of Medicine.
4
Department of Immunology and Microbiology, University of Colorado School of Medicine; Department of Biomedical Research, National Jewish Health; john.cambier@ucdenver.edu.

Abstract

B cells reactive with a specific antigen usually occur at a frequency of <0.05% of lymphocytes. For decades researchers have sought methods to isolate and enrich these rare cells for studies of their phenotype and biology. Approaches are inevitably based on the principle that B cells recognize native antigen by virtue of cell surface receptors that are representative in specificity of antibodies that will eventually be secreted by their differentiated daughters. Perhaps the most obvious approach to the problem involves use of fluorochrome-conjugated antigens in conjunction with fluorescence-activated cell sorting (FACS). However, the utility of these methods is limited by cell frequency and the achievable rate of analysis and isolation by electronic sorting. A novel method to enrich rare antigen-specific B cells using magnetic nanoparticles that results in high yield enrichment of antigen-reactive B cells from large starting cell populations is described. This method enables improved monitoring of the phenotype and biology of antigen reactive cells before and following in vivo antigen encounter, such as after immunization or during development of autoimmunity.

PMID:
28287549
PMCID:
PMC5409333
DOI:
10.3791/55382
[Indexed for MEDLINE]
Free PMC Article

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