title: Optimization and qualification of a memory B-cell ELISpot for the detection of vaccine-induced memory responses in HIV vaccine trials.
abstract: Various aspects of the human immune system can be analyzed to determine the efficacy of a vaccine. We have developed a B-cell ELISpot to measure HIV-specific antibody-secreting B cells in the peripheral blood as a result of vaccination or natural infection. Our method includes stimulating peripheral blood mononuclear cells with interleukin-2 and a polyclonal activator, R848, to induce memory B cells to differentiate into antibody-secreting cells. Total immunoglobulin-secreting as well as antigen-specific B cells are then quantified. We have tested several HIV Env gp120 and gp140 proteins from different HIV subtypes, as well as a sensitive consensus group M Env gp140. Our findings indicate that the B-cell ELISpot provides a sensitive and specific tool to detect antigen-specific memory B-cell responses, and it is equally suited to detect antibody-secreting plasmablasts present in the circulation shortly after infection or vaccination.
authors.affiliation: Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.
authors.forename: Paula N
authors.initials: PN
authors.lastname: Walsh
authors.forename: David P
authors.initials: DP
authors.lastname: Friedrich
authors.forename: Julie A
authors.initials: JA
authors.lastname: Williams
authors.forename: Rebecca J
authors.initials: RJ
authors.lastname: Smith
authors.forename: Terri L
authors.initials: TL
authors.lastname: Stewart
authors.forename: Donald K
authors.initials: DK
authors.lastname: Carter
authors.forename: Hua-Xin
authors.initials: HX
authors.lastname: Liao
authors.forename: M Juliana
authors.initials: MJ
authors.lastname: McElrath
authors.forename: Nicole
authors.initials: N
authors.lastname: Frahm
mesh_terms: D016915:AIDS Vaccines / Q000276:immunology*; D000917:Antibody Formation; D001402:B-Lymphocytes / Q000276:immunology*; D019921:Biotinylation; D002986:Clinical Trials as Topic; D058501:Enzyme-Linked Immunospot Assay / Q000379:methods*; D006801:Humans; D007156:Immunologic Memory
chemical_list: D016915:AIDS Vaccines
keywords: ASC; Ag; Antibody-secreting cells; B-cell ELISpot; HBSS; HIV; Hanks balanced salt solution; Ig; KLH; Keyhole Limpet Hemocyanin; Memory B-cell responses; Mucosal responses; PBMC; PVDF; SFC; TMB; Vaccine; antibody-secreting cell; antigen; immunoglobulin; peripheral blood mononuclear cells; polyvinylidene fluoride; spot-forming cells; tetramethylbenzidine