Immune reconstitution after haplo-HSCT with CD45RA-depleted DLI. (A) CD45RA expression by lymphocytes in the DLI product before and after depletion of CD45RA⁺ cells. A representative DLI product is shown. (B) Schematic overview of DLI administration and blood sample collection. Median days post-transplant are indicated. (C) Experimental approach. Blood samples from patients who received DLI, their donors, and unrelated healthy controls were stained with a flow cytometry panel allowing the detection of innate lymphocytes. The remaining cells of each sample were cultured overnight in presence or absence of a CMV-peptide library, and stained with a T-cell–focused flow cytometry panel. Unstimulated samples were used for bulk T-cell analysis, whereas CMV-specific cells were identified through Boolean gating for activation markers in response to CMV-peptide stimulation. NK cells, bulk, and CMV-specific T-cell phenotypes were analyzed in more detail with unsupervised clustering. In addition, TCR sequencing of blood and DLI samples from patients who received DLI and control patients was performed to assess the impact of the DLI at the clonal level. (D) Median lymphocyte counts with interquartile range in patients who underwent haplo-HSCT and received or did not receive CD45RA-depleted DLI (n = 7-17/group). Statistical significance was determined with Mann-Whitney test. (E) Median lymphocyte counts with interquartile range at T7 in patients who underwent haplo-HSCT and received or did not receive CD45RA-depleted DLI (n = 10-14/group). Statistical significance was determined with Mann-Whitney test. ILC1, group 1 innate lymphoid cell; ILC2, group 2 innate lymphoid cell; ILC3, group 3 innate lymphoid cell; T_CM, central memory T cell; T_EM, effector memory T cell; T_EMRA, effector memory T cell re-expressing CD45RA; T_N, naïve T cell.