CD8⁺ T_eff cells damaged antigen-specific target cells but not bystander cells. (A) Longitudinal fluorescence confocal micrographs (z stacks shown as maximum projection) of islet grafts in ACE, acquired noninvasively in vivo. Visualized were infiltration and damage by OVA-specific GFP-labeled CD8⁺OT1 T_eff cells in OVA⁺CFP⁻ islet grafts (gray) versus bystander OVA⁻CFP⁺ islets (blue). Data represent a total of 6 pairs of antigen-specific OVA⁺CFP⁻ and nonspecific OVA⁻CFP⁺ islets conjoined like Siamese twins (left), total 6 separated OVA⁻CFP⁺ islets (right) in the same ACE environment but not conjoined with OVA⁺CFP⁻ grafts, and total 6 separated OVA⁺CFP⁻ islet grafts, from 5 different mice in three experiments. (B) Changes in relative islet mass (volume; mean ± SEM) of OT1 antigen-specific islets (OVA⁺) compared with bystander CFP islets in nonadjacent positions, after CD8⁺ OT1 T_eff cells transfer. (C) Changes in relative mass (volume; mean ± SEM) of bystander islet grafts conjoined with the specific islet graft or separated from the specific islet graft in the same ACE. Bars, 100 µm. **, P < 0.01; ***, P < 0.001. See Videos 2 and 3 for 3D islet mass visualization before and after CD8⁺ T_eff cell–mediated damage to the specific OVA⁺ tissue.