Analyses of islet β cell proliferation by BrdU incorporation. B6 islets, RIP-mOVA⁺ islets, or a mixture of B6 and RIP-mOVA⁺ islets were transplanted under the kidney capsules of B6 mice. At 7 wk after transplantation, 2 × 10⁶ activated OT1 cells were injected i.v. into the mice. BrdU was administered by i.p injection beginning on day 2 after OT1 T cell transfer. The animals were sacrificed on day 10, and kidney tissue sections were prepared to analyze islet grafts. Tissue sections of islet grafts and surrounding kidney tissues were analyzed by immunostaining with anti-insulin and anti-BrdU antibodies and counterstained for nuclei with DAPI. The RIP-mOVA⁺ islet graft group had a complete destruction of islet grafts, and thus it is not presented. Tissue sections of the B6 islet alone group (A) or B6 and RIP-mOVA⁺ islets mixed engraftment group (B) were compared for insulin⁺ cells with BrdU signals. A serial section of the latter group was also used to show the secondary staining only (C). Inset in B is the zoomed-in image of the highlighted area. The BrdU⁺ Insulin⁻ cells were likely the infiltrating lymphocytes that destroyed the RIP-mOVA⁺ islets in the mixed engraftment group. (D) Data were pooled from 2 experiments with 4 mice, with 36 and 34 islets analyzed in the B6 islet alone engraftment and the mixed engraftment groups, respectively. Each data point represents one islet graft (mean ± SEM). **, P < 0.01. Bar, 50 µm.