Isolation and characterization of human antigen-specific B lymphocytes.
Antigen-specific B cells from the peripheral blood of immunized donors were isolated following rosette formation with antigen-coated immunomagnetic beads and were phenotypically and functionally characterized. B cells separated with tetanus toxin (TT)- or keyhole limpet hemocyanin (KLH)-coated beads produced significant amounts of specific IgM and IgG antibodies when they were transformed with Epstein-Barr virus or cultured with autologous T cells in the presence of a specific antigen. Cells isolated from peripheral blood mononuclear cells (PBMC) with TT-coated beads consisted mostly of B cells (88.7% CD20+). B cells detected among the rosetting cells were predominantly sIgM+ with low percentages of the other isotype-expressing B cells. The mean percentage of TT-specific B cells among adult donors was 0.34% of PBMC, and increased to up to 2% following a booster immunization with antigen. A similar increase in the number of rosetting B cells was also observed following in vitro culture of PBMC with antigen. These findings demonstrate that immunomagnetic bead selection serves as a specific and reliable approach in identifying and isolating antigen-specific B cells from human PBMC and provides a valuable method for studying T-B interactions in antigen-specific immune responses.
Oshiba
A
A
Department of Pediatrics, National Jewish Center for Immunology and Respiratory Medicine, Denver, Colorado 80206.
Renz
H
H
Yata
J
J
Gelfand
E W
EW
D000328:Adult; D001402:B-Lymphocytes / Q000276:immunology*; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D005434:Flow Cytometry; D006433:Hemocyanins / Q000276:immunology*; D004854:Herpesvirus 4, Human / Q000502:physiology; D006801:Humans; D007132:Immunoglobulin Isotypes / Q000096:biosynthesis; D018189:Immunomagnetic Separation / Q000379:methods*; D016130:Immunophenotyping; D012397:Rosette Formation / Q000379:methods; D013601:T-Lymphocytes / Q000276:immunology; D013744:Tetanus Toxin / Q000276:immunology*
D000939:Epitopes; D007132:Immunoglobulin Isotypes; D013744:Tetanus Toxin; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin