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. 2014 Oct 22;9(10):e110883. doi: 10.1371/journal.pone.0110883

Figure 1. Evaluation and characterization of immediate effector T cell responses induced by a single immunization with the CAL-SHIV-IN− DNA vaccine.

Figure 1

A) PBMCs isolated from blood samples taken at the indicated weeks post-immunization (PI) were used for ELISPOT assay to detect IFN-γ producing cells in response to the pools of SIV Gag (indicated in blue), HIV Env (indicated in red), HIV Tat+Rev+ SIV Nef (TRN, indicated in green) and SIV Pol (indicated in purple) peptides. Numbers of IFN-γ producing cells obtained per million of PBMCs against all Ags tested are indicated in the y-axis. B) Mean and range of IFN-γ ELISPOT responses to Gag and TRN pools across 22 (for BX72, 78, 80, 83, 84) and 14 (for BX73) time-points collected. C) Memory phenotyping of Gag-specific T cells at W8 PI. PBMCs were restimulated for 16 h in presence or absence of Gag pool of peptides in medium containing Brefeldin A. Cells were then surface stained with EMA, CD3, CD8, CD4, CD28, CD95 mAbs, permeabilized and stained with IFN-γ, IL-2 and TNF-α mAbs. For analysis, cells were initially gated on live lymphocytes (low FSC/SSC, EMA-, CD3+), CD8 bright T cells (orange color) and CD4+ T cell populations (blue color). Antigen-specific T cells were identified by their capacity to secrete one or more cytokines (black dots). For each indicated animal (BX72, 78, 80, 84), we displayed the maximal response obtained at this time point within CD8+ or CD4+ T cell populations. For simplicity, the percentage of total responses (total black dots) obtained through all quadrants (IFN-γ+, IFN-γ+ &TNF-α+, IFN-γ+&IL-2+ and IL-2+) are indicated after the subtraction of background obtained with cells cultured with medium only. For memory T cell subset determinations, all black dots were superimposed to the total CD8+ or CD4+ T cell populations and plotted against memory markers (CD28 and CD95). The naïve population was defined as CD28+ CD95−, effector memory (EM) as CD28− CD95+ and central memory (CM) as CD28+ CD95+. D) Sequential evaluation of SIV Gag p55 specific antibody responses in immunized macaques (“homemade kit”). E) Sequential evaluation of HIV Env gp160 specific antibody responses in immunized macaques (Biorad, France).