title: Fluorescently labeled dengue viruses as probes to identify antigen-specific memory B cells by multiparametric flow cytometry.
abstract: Low frequencies of memory B cells in the peripheral blood make it challenging to measure the functional and phenotypic characteristics of this antigen experienced subset of B cells without in vitro culture. To date, reagents are lacking to measure ex vivo frequencies of dengue virus (DENV)-specific memory B cells. We wanted to explore the possibility of using fluorescently labeled DENV as probes to detect antigen-specific memory B cells in the peripheral blood of DENV immune individuals. Alexa Fluor dye-labeled DENV yielded viable virus that could be stored at -80°C for long periods of time. Using a careful gating strategy and methods to decrease non-specific binding, we were able to identify a small frequency of B cells from dengue immune individuals that bound labeled DENV. Sorted DENV(+) B cells from immune, but not naïve donors secreted antibodies that bound DENV after in vitro stimulation. Overall, Alexa Fluor dye-labeled DENVs are useful reagents to enable the detection and characterization of memory B cells in DENV immune individuals.
authors.affiliation: Division of Infectious Disease and Immunology, University of Massachusetts Medical School, Worcester, MA 01655, USA.
authors.forename: Marcia
authors.initials: M
authors.lastname: Woda
authors.affiliation: Division of Infectious Disease and Immunology, University of Massachusetts Medical School, Worcester, MA 01655, USA. Electronic address: anuja.mathew@umassmed.edu.
authors.forename: Anuja
authors.initials: A
authors.lastname: Mathew
mesh_terms: D000914:Antibodies, Viral / Q000276:immunology; D000956:Antigens, Viral / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D002460:Cell Line; D003716:Dengue Virus / Q000276:immunology*; D005434:Flow Cytometry / Q000379:methods; D005453:Fluorescence; D006801:Humans; D007156:Immunologic Memory / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral
keywords: B cells; Dengue; Flow cytometry; Labeled viruses