Cellular immune responses induced by the DNA vaccine candidates.

<p>Intra-cellular INF gamma (IFNγ) staining IFNγ production was measured by FACS analysis after intracellular staining of IFNγ). Balb/C mice (n = 6 per group) were inoculated IM with (i) pORT ConcTat NG, (ii) pORT Gag 203 , (iii) pORT Env NG2 , (iv) pORT Env NG9 , (v) pORT gp120 C N54 (positive control), (vi) pORT1a clade (negative control). One week after the second/third booster immunization, spleen cells were isolated and tested for specific cellular immune responses by measuring IFN-g production after stimulation with pools of overlapping 15-mer peptides. Shown is the total number of CD8⁺ IFNγ⁺ cells per 30.000 CD8⁺ restimulated mouse spleen cells. A) mice 1–3, B) mice 4–6, C) merged data.</p

Neutralizing antibody titers of plasmas tested with autologous and heterologous viral envelopes.

<p>Plasma samples are from chronically infected individuals not currently on treatment. Titers are expressed as 1/plasma dilution. Higher numbers indicate greater inhibition (titer). Green color indicates positive neutralization activity which is significantly greater than inhibition of the specificity control, aMLV. Dashed red line indicates autologous plasma/virus pair. na = not available.</p

Viruses and plasmas used in antibody neutralization assays.

<p>Viruses and plasmas used in antibody neutralization assays.</p

The IC₅₀ values of anti-V3 mAbs against 57 HIV-1 pseudoviruses tested using the U87 target cell line.

<p>The IC₅₀ values were estimated from the titration curves of all mAb/psV combinations and are highlighted according to the color-coded scale. Pseudoviruses expressing Envs of JRCSF, NL3.4, and SF162 were tested as positive controls, whereas the irrelevant anti-parvovirus mAb 860 and aMLV Env-expressing psV were used as negative controls. When 50% neutralization was not achieved at the highest mAb concentration tested (50 µg/ml), the IC₅₀ values are shown as >50.</p

The neutralization of nine HIV-1 pseudoviruses by mAb 2191 using U87 as target cells.

<p>The neutralization curves of anti-V3 mAb 2191 against nine selected HIV-1 pseudoviruses are shown with their corresponding AUC and IC₅₀ values. Fifty percent neutralization is denoted by the dashed line. Significant neutralization at the confidence level of p<0.001 (denoted with * after AUC values) was determined statistically based on comparison with the AUC values of the negative controls together with the slopes of the titration curves as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0010254#s2" target="_blank">Materials and Methods</a>. For viruses coming from patients where the date of infection is known, viruses are denoted as coming from acutely- or chronically-infected patients, and the clade of the virus is denoted by the capital letter in its name (A, B, C or D).</p

Neutralization curves of anti-V3 mAbs against representative HIV-1 psVs tested using the TZM.bl target cell line.

<p>Seven anti-V3 mAbs were tested against HIV-1 psVs in the TZM.bl experiment. The titration curves observed against nine selected viruses are shown. The background neutralization observed with the irrelevant control mAb 860 is also shown for comparison. Curve fitting was performed using the polynomial regression (quadratic) model described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0010254#s2" target="_blank">Materials and Methods</a>. Fifty percent neutralization is denoted by the dashed line.</p

The AUC values of anti-V3 mAbs against 57 HIV-1 pseudoviruses tested using the U87 target cell line.

<p>AUC values were estimated from the titration curves as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0010254#s2" target="_blank">Materials and Methods</a> section. Statistically significant neutralization at p<0.001 is color-coded according to the designated scale.</p

Characteristics of human mAbs used for this study.

<p>nd – not determined.</p><p>PA – protective antigen of anthrax.</p><p>§– Gorny et al., unpublished data.</p

Neutralization curves of anti-V3 mAbs against representative HIV-1 psVs tested using the U87 target cell.

<p>Seven anti-V3 mAbs (2191, 2219, 2557, 2558, 3074, 3869, and 447) were tested for neutralization against HIV-1 pseudoviruses using the U87 cell line as target cells. MAb 860, specific for parvovirus, and a pseudovirus expressing the aMLV Env were used as negative controls in this assay. The titration curves from eight selected psVs and the aMLV psV control are shown. Curve fitting was performed using the polynomial regression (quadratic) model described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0010254#s2" target="_blank">Materials and Methods</a>. Fifty percent neutralization is denoted by the dashed line. Virus nomenclature is denoted as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0010254#pone-0010254-g001" target="_blank">Figure 1</a>.</p

The AUC values of anti-V3 mAbs against 41 HIV-1 pseudoviruses tested using the TZM.bl target cell line.

<p>The AUC values were calculated from the titration curves as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0010254#s2" target="_blank">Materials and Methods</a> section. All mAb/psV pairs with statistically significant neutralization at p<0.001 are color-coded according to the designated scale. The irrelevant anti-parvovirus mAb 1418 was used together with three other control mAbs (data not shown) as negative controls.</p