CD28- and CTLA-4–mediated signals elicit opposite effects on tissue infiltration by T cells. Female and male CBA/Ca mice were treated intraperitoneally with 600 U IFNγ. After 48 hours, 5 × 106 PKH26-labeled HY-specific CD8+ C6 T cells were injected intraperitoneally in PBS (A-B). In some mice, T cells were coinjected with a mixture of hamster anti–mouse CD28 (5 μg/5 × 106 cells) and rabbit anti–hamster Ig (2.5 μg/5 × 106 cells; C,I) or hamster anti–mouse CD152 (2.5 μg/5 × 106 cells) and rabbit anti–hamster Ig (1.25 μg/5 × 106 cells; D,J) together with the PKH26-labeled C6 cells. As a control, mice were injected with a mixture of hamster Ig (5 μg/5 × 106 cells, the highest Ig dose used for cross-linking) and rabbit anti–hamster Ig (2.5 μg/5 × 106 cells) together with labeled T cells (B,H). The presence of labeled T cells in the peritoneal membrane and lavage was analyzed after 24 hours by wide-field fluorescence microscopy and flow cytometry, respectively. To minimize the effect of arbitrary choice of field, × 10 magnifications are shown. Tissue infiltration was quantified by randomly selecting ten × 10-magnified fields and assessing the number of fluorescent cells in each field. The mean T-cell infiltration ± SD observed in samples from at least 3 animals are summarized in panels E,K (infiltration of the peritoneal membrane) and F,L (cells retrieved in the peritoneal lavage). E, *P < .004; F, *P < .007; K, *P < .05 for CD28 and *P < .001 for CD152; L, *P < .02 for CD28 and *P < .001 for CD152.
