Anti–CD19-CAR–transduced T cells can be detected in the blood of patients for up to 181 days after infusion. (A) The percentage of total PBMCs that contained the anti-CD19 CAR gene was determined by quantitative PCR. In patients 1 through 6, the peak levels and persistence of cells containing the CAR gene varied. (B) The percentage of total PBMCs from patient 7 and patient 8 that contained the anti-CD19 CAR gene was determined by quantitative PCR. Compared with the other patients, these patients had substantially greater persistence of cells that contained the CAR gene. (C) Anti-CD19 CAR expression was detected ex vivo on PBMCs of patient 7 by flow cytometry staining with labeled CD19 protein (CD19-DDK) 13 days after CAR-transduced T-cell infusion. Only background levels of cells expressing the anti-CD19 CAR were detected before treatment. (D) In patient 8, T cells expressing the anti-CD19 CAR were detected by flow cytometry staining with labeled CD19 protein 13 days after CAR-transduced T-cell infusion. Only background levels of cells expressing the anti-CD19 CAR were detected before treatment. (C-D) The plots are gated on CD3⁺ lymphocytes. (E) PBMCs from patient 7 were cultured for 4 hours with either CD19⁺ CD19-K562 cells or NGFR-K562 control cells that do not express CD19. Before treatment, CD107a was not up-regulated on T cells after culture with either CD19-K562 cells or NGFR-K562 cells. In contrast, T cells from 8 days after infusion up-regulated CD107a after a 4-hour culture with CD19-K562 cells but not NGFR-K562 cells. All plots are gated on CD3⁺ lymphocytes.