Comparison of quantitation of the absolute numbers per 10⁵ T cells of anti-EBV and alloreactive T cells by LDA and by intracellular IFN-γ staining over the course of culture.

(A, C) Absolute numbers of anti-EBV–reactive CD3⁺ cells per 10⁵ T cells by LDA (♦) and absolute numbers of intracellular IFN-γ–producing CD3⁺ (▪), CD8⁺ (▵), and CD4⁺ (○) per 10⁵T cells in response to stimulation with autologous EBV BLCL of donor A and donor B, respectively. (B, D) Absolute numbers of anti-alloreactive CD3⁺ cells per 10⁵ T cells by LDA (♦) and the absolute numbers of intracellular IFN-γ–producing CD3⁺(▪), CD8⁺ (▵), and CD4⁺ (○) per 10⁵ T cells in response to stimulation with allogeneic EBV BLCL of donor A and donor B, respectively. Despite the 25- to 90-fold higher quantitation of EBV-specific and the 30- to 220-fold higher quantitation of alloreactive T-cell numbers obtained by determination of intracellular IFN-γ staining, the alterations in frequencies detectable by both assays largely paralleled each other over the course of culture.