Characterization of the CD19-CAR_Lenti T cells at the end of manufacturing (DPs) and after infusion. The DPs were characterized in terms of number of total cells obtained at the end of manufacturing (A); levels of transduction (B); vector copy number per cell (C); CD4⁺ and CD8⁺ subpopulations (D); and memory phenotype, with identification of naïve, CM, EM, and TEMRA CD45RA cells in CD4⁺ and CD8⁺ T cells (E). After infusion, the peak levels of circulating CAR T cells per μL in the PB were evaluated by flow cytometry for each DL (F) and the expansion of CD19-CAR__Lenti T cells was monitored overtime by both qPCR (G) and flow cytometry, the latter identifying the absolute numbers of circulating CAR T cells (H) and the percentage of CD3 cells expressing the CAR molecule (I). The CD4/CD8 phenotype of circulating CD19-CAR_Lenti T cells was also evaluated by flow cytometry in the PB, both in terms of CD4/CD8 distribution (J) and of memory phenotype of each subpopulation (K-L). Homing to the BM and persistence of CD19-CAR_Lenti T cells was analyzed by flow cytometry and qPCR (M-N). (O) CD4/CD8 phenotype of BM infiltrating CD19-CAR_Lenti T cells, evaluated by flow cytometry. N, naïve; Pb, peripheral blood.