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1.
Figure 3.

Figure 3. From: Transmembrane Domain Membrane Proximal External Region but Not Surface Unit-Directed Broadly Neutralizing HIV-1 Antibodies Can Restrict Dendritic Cell-Mediated HIV-1 Trans-infection.

Both mature dendritic cell (mDC)–mediated trans-infection and cell-free infection are similarly inhibited by b12 antigen-binding fragment (Fab). The x-axis shows the amount of b12 immunoglobulin G (IgG) and b12 Fab used in log μg/mL, and the y-axis shows the percentage inhibition relative to infection without any inhibitor. Each point represents an average of at least 3 independent experiments performed in triplicate and is the mean percentage inhibition, with whiskers denoting standard errors of the mean. Nonlinear regression was used to estimate a fitted curve in GraphPad Prism 5.

Manish Sagar, et al. J Infect Dis. 2012 April 15;205(8):1248-1257.
2.
Figure 1.

Figure 1. From: Transmembrane Domain Membrane Proximal External Region but Not Surface Unit-Directed Broadly Neutralizing HIV-1 Antibodies Can Restrict Dendritic Cell-Mediated HIV-1 Trans-infection.

Target cells were exposed to similar amounts of infectious virus when incubated with mature dendritic cell (mDC)–laden human immunodeficiency virus type 1 (HIV-1) vs cell-free stocks. The x-axis identifies the box plots for the relative light units (RLUs) 2 d after TZM-bl infections among the various HIV-1 isolates in mDC (unfilled box) vs cell-free (gray boxes) infections. The black line in each box represents the median value, boxes show the interquartile range, and whiskers show the maximum and minimum values. Values were generated from a minimum of 10 independent experiments with at least 2 different virus stocks for each isolate.

Manish Sagar, et al. J Infect Dis. 2012 April 15;205(8):1248-1257.
3.
Figure 6.

Figure 6. From: Transmembrane Domain Membrane Proximal External Region but Not Surface Unit-Directed Broadly Neutralizing HIV-1 Antibodies Can Restrict Dendritic Cell-Mediated HIV-1 Trans-infection.

Anti-gp41 broadly neutralizing antibody 4E10 accumulates at the mDC–T cell synaptic junction in a human immunodeficiency virus (HIV)–independent manner. Mature DCs were incubated with 4E10 (A and C) or 2G12 (B and D) prior to coculture with autologous CD4+ T cells prestained with a blue fluorescent dye. Mature DC–T cell synaptic junctions were visualized by staining with anti-CD81 monoclonal antibodies 4E10 (red) and 2G12 (red) (A and B; middle panels). Alternatively, mature DCs preloaded with Gag-eGFP virus-like particles (green) were incubated with 4E10 (C; red) or 2G12 (D; red) and cocultured with autologous CD4+ T cells. Representative deconvolution images of mDC–T cell cocultures (×100 magnification) are shown. Merged bright field images for each of the stains are shown in the right panels.

Manish Sagar, et al. J Infect Dis. 2012 April 15;205(8):1248-1257.
4.
Figure 2.

Figure 2. From: Transmembrane Domain Membrane Proximal External Region but Not Surface Unit-Directed Broadly Neutralizing HIV-1 Antibodies Can Restrict Dendritic Cell-Mediated HIV-1 Trans-infection.

Neutralization of mature dendritic cell (mDC)–mediated human immunodeficiency virus trans-infection by anti-gp120–directed broadly neutralizing antibodies is attenuated compared with cell-free virus infection. Mature DC–associated (filled squares) and cell-free (hollow squares) neutralization is compared for Q23 (A and F), YU-2 (B and G), JRCSF (C and H), REJO (D and I), and CH077 (E and J). The x-axis shows the amount of VRC01 (AE) and PG16 (FJ) used in log μg/mL, and the y-axis shows the percentage inhibition relative to infection without any antibody. Each point represents an average of at least 3 independent experiments performed in triplicate and is the mean percentage inhibition, with whiskers denoting standard errors of the mean. Nonlinear regression was used to estimate a fitted curve in GraphPad Prism 5.

Manish Sagar, et al. J Infect Dis. 2012 April 15;205(8):1248-1257.
5.
Figure 4.

Figure 4. From: Transmembrane Domain Membrane Proximal External Region but Not Surface Unit-Directed Broadly Neutralizing HIV-1 Antibodies Can Restrict Dendritic Cell-Mediated HIV-1 Trans-infection.

Mature dendritic cell (mDC)–mediated transfer and cell-free infection are equally susceptible to anti-gp41–directed broadly neutralizing antibodies. Neutralization for Lai (A and E), Lai/Balenv (B and F), JRCSF (C), NL4-3 (D and H), and 89.6 (G) is compared among mDC-associated virus (filled squares), cell-free virus (hollow squares), and mDC-associated virus preincubated with FcR blocking reagent (filled circles). The x-axis shows the amount of 2F5 (AD) and 4E10 (EH) used in log μg/mL, and the y-axis shows the percentage inhibition relative to infection without any antibody. Each point represents an average of at least 3 independent experiments performed in triplicate and is the mean percentage inhibition, with whiskers denoting standard errors of the mean. Nonlinear regression was used to estimate a fitted curve in GraphPad Prism 5.

Manish Sagar, et al. J Infect Dis. 2012 April 15;205(8):1248-1257.
6.
Figure 5.

Figure 5. From: Transmembrane Domain Membrane Proximal External Region but Not Surface Unit-Directed Broadly Neutralizing HIV-1 Antibodies Can Restrict Dendritic Cell-Mediated HIV-1 Trans-infection.

Anti-gp41–specific broadly neutralizing antibodies (bNAbs) bind the surface of mature dendritic cells (mDCs) in the absence of human immunodeficiency virus type 1 (HIV-1) particles. Mature DCs were incubated with 4E10, 2F5, 2G12, b12, PG16, VRC01, or phosphate-buffered saline alone and were then washed and stained for human immunoglobulin G (IgG), using Alexa 594–conjugated secondary antibody and for nucleus using DAPI. Representative deconvolution merged bright field images (×100 magnification) of mDC staining observed with 4E10 (A) and 2G12 (B) bNAbs is shown. Arrowheads indicate IgG binding to the mDC surface. C, The y-axis shows the percentage of mDCs bound by various bNAbs, shown on the x-axis. The black line in each box represents the median value; boxes show the interquartile range, and whiskers show the maximum and minimum values. The background signals were calculated as a mean plus 2 times the standard deviation of the signals from cells without bNAb. Cells expressing a signal above the background on the cell membrane were counted as positive for IgG binding. Graphs were derived from counting at least 10 images from each sample, with a minimum of 100 cells in 3 independent experiments with cells derived from 3 different donors.

Manish Sagar, et al. J Infect Dis. 2012 April 15;205(8):1248-1257.

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