CD1d/GPI dimer staining of T cells. (A) Representative flow cytometry plots showing ex vivo staining of patient and control PBMCs with anti-CD8 and CD1d/t-GPI dimer followed by analysis by multiparameter flow cytometry. Plots shown are gated on DAPI⁻CD14⁻CD19⁻CD3⁺ cells. (B) Cumulative data showing frequency of CD8⁺CD1d/t-GPI dimer⁺ T cells from 4 PNH patients and 4 healthy controls after ex vivo staining of PBMCs as described in panel A (staining with CD1d/Veh dimer was not performed in this set of experiments). (C) Representative flow cytometry plots showing staining of patient CD8⁺ T cells with (left) CD1d/t-GPI or (right) unloaded dimer after 14 days of coculture with C1R-CD1d cells loaded with t-GPI. Gating strategy as described in panel A. (D) Cumulative data showing frequency of CD8⁺ CD1d/t-GPI dimer⁺ vs CD1d/Veh dimer⁺ T cells from patients (n = 7) and controls (n = 11), after 14 days of coculture of CD8⁺ T cells with C1R-CD1d cells loaded with t-GPI (1-way analysis of variance with Tukey multiple comparison test; NS, not significant). (E) Staining of CD8⁺ T cells with CD1d/h-GPI dimer or CD1d/Veh dimer after 14 days of coculture with C1R-CD1d cells loaded with h-GPI. Gating strategy as described in panel A. Staining of T cells from a PNH patient is shown. (F) Cumulative data from 10 PNH patients and 7 normal controls.