Naturally occurring human CD4⁺CD25⁺ cells can be found in healthy individuals and are phenotypically distinct from recently activated CD4⁺ cells.

(A) CD4⁺CD25⁺ cells constitute a significant proportion of CD4⁺ T cells in peripheral blood of healthy individuals. Purified CD4⁺ T cells were stained with anti-CD4 FITC and anti-CD25 PE. The resultant purity of CD4⁺CD25⁺ and CD4⁺CD25⁻ cells isolated using magnetic beads from a typical experiment is shown. (B) Phenotype of CD4⁺CD25⁺ cells from peripheral blood of healthy individuals. In the top panel, 100 μL peripheral blood was stained with anti-CD4–QR, anti-CD25⁻ PE, and FITC-conjugated monoclonal antibodies as indicated. The dot plots were gated on CD4⁺ lymphocytes. In the bottom panel, 100 μL peripheral blood was stained with anti-CD4–QR, anti-CD25⁻FITC, and anti–CTLA-4–PE (thin solid lines) or isotype-matched control (thick solid lines). The histogram plot was gated on a CD4⁺CD25⁺ or CD4⁺CD25⁻lymphocyte population. (C) The phenotype of recently activated CD4⁺ cells that express CD25 is distinct from “constitutive” CD4⁺CD25⁺ cells. PBMCs from the same healthy individuals were stimulated with PHA (2 μg/mL) for 2 days. The cells were harvested and stained with anti-CD4–QR, anti-CD25⁻ PE, and the indicated FITC-conjugated monoclonal antibodies. The dot plots were gated on CD4⁺lymphocytes.