Figure 2.

CD8-NPs are specific toward CD8⁺ T cells

(A) Flow cytometry gating strategy for determining specificity of CD8-NPs for CD8⁺ T cells. Cells were first gated based on the CD3⁺ lymphocyte population (left). The unstained sample was then used to determine the quadrant gate placement for the CD4 stain and NP fluorescence (middle). A representative plot for the distribution of CD8-NPs in CD4^– and CD4⁺ T cells is shown on the right. Quadrant 1 represents the CD4^– T cells that are positive for CD8-NPs. Quadrant 2 represents the CD4⁺ T cells that are positive for CD8-NPs. Quadrant 3 represents CD4⁺ T cells that are negative for CD8-NPs. Quadrant 4 represents CD4^– T cells that are negative for CD8-NPs. Representative plots for n = 4 independent experiments. (B) Distribution of CD8-NPs in HD CD4^– and CD4⁺ T cells within a CD3⁺ T cell population (the percent of CD8-NPs bound to CD4^– T cells versus the percent of CD8-NPs bound to the CD4⁺ T cells; n = 4). (C) Percentage of HD CD4^– and CD4⁺ T cells within a CD3⁺ T cell population that are positive for CD8-NPs (n = 4). (B and C) Data are bar graphs representing the mean ± standard deviation. Comparisons are made using paired Student’s t test. (D) Flow cytometry gating strategy for determining non-specific binding of NPs (labeled with IgG antibody) for CD8⁺ T cells. Cells were first gated based on the CD3⁺ lymphocyte population (left). The unstained sample was then used to determine the quadrant gate placement for the CD4 stain and NP fluorescence (middle). A representative plot for the distribution of IgG-NPs in CD4^– and CD4⁺ T cells is shown on the right. Quadrant 1 represents the CD4^– T cells that are positive for IgG-NPs. Quadrant 2 represents the CD4⁺ T cells that are positive for IgG-NPs. Quadrant 3 represents CD4⁺ T cells that are negative for IgG-NPs. Quadrant 4 represents CD4^– T cells that are negative for IgG-NPs. Representative plots for n = 3 independent experiments.