Resistance of cord blood T_regs toward CD95L-mediated cell death. (A) FACS-sorted adult T_regs (CD4⁺CD25^hi), cord blood T_regs (CD4⁺CD25⁺), and their respective T_convs were incubated with CD95L (1:20) for 10 hours, and specific cell death rates were determined by FACS analysis as described previously.¹²  Similarly, CD4⁺CD25⁺CD45RO^lo T cells (R2) as well as CD4⁺CD25⁺CD45RO^hi T cells (R1) were FACS sorted as shown in panel B, and all cells were incubated with CD95L (1:20) for 10 hours. Error bars represent SEM. (B) Enrichment of naive or memory cord blood T_regs by FACS sorting. Cord blood lymphocytes were stained with fluorochrome-labeled mAbs against CD4, CD25 and CD45RO. Previous to FACS sorting, one aliquot from the sort sample was additionally stained with anti-CD95 mAb to detect CD4⁺CD25⁺CD95^hi T cells. Because CD4⁺CD25⁺CD95^hi T cells cosegregate with CD45RO^hi T cells (arrow), the sorting gate was set on CD45RO^hi cells to separate CD4⁺CD25⁺CD45RO^hi CD95^hi memory T cells in R1 from the naive cord blood T_regs in R2. See the penultimate paragraph of “Results” for further explanation. One representative experiment of at least 3 independent experiments is shown.