Summary of analyses for percentage of CD25⁺ cells among TCR⁺CD4⁺ cells by flow cytometry. Thymocytes (A), splenocytes (B), portal (C), or inguinal lymph node cells (D) from DO11.10 mice were analyzed 30 or 60 days after ova (•) or GFP (▪) gene transfer (n = 4 per time point and group). Anti-CD4 (conjugated to PE-Cy5.5), anti-CD25 (PE), and KJ1-26 (FITC) antibodies were used for staining. Initial gating of viable lymphocytes was performed to identify CD4⁺ cells. CD4⁺ cell population was subsequently used to determine percent TCR⁺CD25⁺ cells. Error bars represent SD except for portal nodes, which had been pooled prior to flow cytometry. Examples for flow cytometry contour plots (inguinal lymph nodes, day 60) are shown in panels E (AAV-EF1α-GFP) and F (AAV-EF1α-ova). Percent TCR⁺CD4⁺CD25⁺ of TCR⁺CD4⁺ is indicated in top right quadrant.