Skip to main content
. 2015 Apr 24;12(2):2443–2450. doi: 10.3892/mmr.2015.3685

Figure 5.

Figure 5

Cytotoxicity of mWT1-specific CTLs induced by caTLR4-PMDC11. mWT1-specific cytotoxicity of CTLs generated from normal CD8+ T cells by priming with mWT1 peptide-pulsed PMDC11, LPS-stimulated PMDC11 or doxycycline-exposed caTLR4-PMDC11 cells was evaluated using GFP-transduced T2A24 cells pulsed with or without mWT1 peptide as target cells by flow cytometric analysis. Target cells were cultured with effector cells in RPMI 1640 without phenol red for 4 h at 37°C. Co-cultured cells were stained with 7AAD and subjected to flow cytometric analysis. Cells were acquired for 120 sec and viable target cells (GFP+/7AAD−) were counted. The cytotoxicity was calculated using the following formula: [(absolute number of viable target cells in the tube containing target cells only - absolute number of viable target cells in the sample tube)/absolute number of viable target cells in the tube containing target cells only] ×100%. (A) Flow cytometry-based cytotoxicity assay of CD8+ T cells cultured with three types of PMDC11 cells. Gated cells in the dot plots represent the number of viable target cells; two repetitions of the experiment produced similar results. (B) Cytotoxicity of CD8+ T cells cultured with three types of PMDC11 cells against mWT1 peptide-pulsed T2A24 cells. (C) Cytotoxicity of CD8+ T cells cultured with caTLR4-PMDC11 cells against T2A24 cells pulsed with or without mWT1 peptide. Statistical differences between the two experimental groups were evaluated by two-way analysis of variance. The values are expressed as the mean ± standard error of the mean. APC, antigen-presenting cell; CTL, cytotoxic T lymphocyte; LPS, lipopolysaccharide; GFP, green fluorescent protein; 7AAD, 7-aminoactinomycin D.