Fig. 4
Characterization of TCRαβ/CD19-depleted HSPC grafts. Apheresis product was either processed immediately after apheresis (fresh mob LP, red) or after storage at 2–6 °C for 48 h (aged mob LP, patterned red). The TCRαβ/CD19-depleted product (fresh TCB2, blue) was determined immediately after processing of fresh or aged mob LP or after storage at 2–6 °C for 48 h (aged TCB2, light blue). A Relative contribution of individual mature leukocyte subpopulations in starting material and target cell fractions among CD45+ leukocytes at the different time-points. The percentage of each subtype was normalized to 100% CD45+ leukocytes. B Percentage of viable CD45+7-AAD− leukocytes in all fractions at different time-points. The efficiency of depletion of TCRαβ+ T cells and CD19+ B cells was compared for products C derived from mobilized apheresis product (mob LP) processed immediately after apheresis (fresh mob LP) or after storage at 2–6 °C for 48 h (aged mob LP) and D derived from processes using NS or LS depletion reagents, respectively. Log depletion for each individual run is depicted with dots and squares representing products generated from fresh and aged mob LP or from NS and LS reagents, respectively. Red line indicates mean values achieving mean depletion efficiency of around 4 log in all tested settings. Box plots and whiskers represent median with interquartile range and min and max values of all 10 depletion runs, respectively. Mean yield of CD34+ stem cells, NK cells and TCRγδ+ cells among total CD45+ cells in depleted products (E) derived from fresh or aged mob LP and F from processes using NS or LS depletion reagents showing a good recovery of cells of interest. G A CFU-C assay was performed via a 14-day culture followed by counting total number of colonies of mob LP and TCB2 at different time-points. Each individual run is plotted, as is mean ± SEM. H The residual alloreactive TCRαβ+ T cell number in HSPC graft was calculated per 4 × 106 CD34+ stem cells representing the transplant dosage of 4 × 106 CD34+ stem cells per kg as the main active component. All data were tested for statistical significance of differences using Student’s t-test. Shown are individual runs and mean ± SEM, n = 4–10 individual runs; ns (not significant) indicates a p > 0.05, *: p < 0.05, **: p < 0.01, ***: p < 0.001. P-values are reported above the corresponding groups