CD4⁺CD25⁺ T cells from B-cell NHL inhibit cytokine production of infiltrating CD4⁺CD25^- T cells. (A) A representative ELISPOT image of IFN-γ and IL-4 secretion by an unsorted cell suspension and by CD4⁺ T cells isolated from biopsy specimens of B-cell NHL. (B) Bar graph showing the average number (± SE; n = 6) of IFN-γ and IL-4 spot-forming cells (SFCs) produced by 5 × 10⁵ unsorted, CD4⁺ or non-CD4⁺ cells on PHA stimulation. (C) IFN-γ and IL-4 production by CD4⁺CD25^- T cells determined by intracellular staining. IFN-γ- or IL-4-positive cells were determined by gating on the corresponding isotype control. (D) Bar graph showing the average number (± SE; n = 6) of IFN-γ and IL-4 SFCs produced by 5 × 10⁵ total CD4⁺CD25^- T cells incubated with or without CD4⁺CD25⁺ T cells on PHA stimulation. Significantly greater IFN-γ secretion (^*P < .05) and IL-4 secretion (#P < .01) was seen when compared with cells incubated in the presence of T_reg cells. Patient sample nos. 5 to 10 were used in these experiments.