Bremm Melanie, Cappel Claudia, Erben Stephanie, Jarisch Andrea, Schumm Michael, Arendt Andreas, Bonig Halvard, Klingebiel Thomas, Koehl Ulrike, Bader Peter, Huenecke Sabine
Clinic for Pediatric and Adolescent Medicine, University Hospital, Frankfurt, Germany.
Department of Hematology/Oncology, Children's University Hospital, Tübingen, Germany.
Cytometry B Clin Cytom. 2017 Mar;92(2):126-135. doi: 10.1002/cyto.b.21328. Epub 2015 Dec 14.
The depletion of TCRαβ T cells and CD19 B cells is a graft purification method for haploidentical stem cell transplantation (HSCT) retaining stem cells, NK cells and TCRγδ T cells. To avoid treatment-related occurrence of severe GvHD a precise quantification of residual TCRαβ T cells in the graft is of essential importance.
Nine stem cell grafts were purified immunomagnetically on a CliniMACS device and flow cytometric quality control (QC) was performed before and after TCRαβ/CD19-depletion.
As a challenge a new 10-color QC-panel was established, which enables accurate quantification of the graft composition. The binding sites of residual TCRαβ T and CD19 B cells were at least partly occupied by depletion antibodies impeding flow cytometric analysis. Based on respective controls and an assumed variation coefficient of 18%, the detection limit of residual TCRαβ T cells was 1 cell/µl. and 0.002% of CD45 cells. Log-depletion of TCRαβ and CD19 cells was -3.9 and -3.3, respectively. The recovery was 82.1%, 67.1% and 72.7% for stem cells, NK cells and for TCRγδ T cells.
In clinical use this method may help to improve transplantation outcome, due to the correct application of the desired stem cell and the limited T cell dose. The panel is designed for the QC following TCRαβ/CD19-depletion but is adaptable to other depletion strategies as well. © 2015 International Clinical Cytometry Society.
TCRαβ T细胞和CD19 B细胞的清除是一种单倍体同基因干细胞移植（HSCT）的移植物纯化方法，可保留干细胞、自然杀伤（NK）细胞和TCRγδ T细胞。为避免与治疗相关的严重移植物抗宿主病（GvHD）的发生，精确量化移植物中残留的TCRαβ T细胞至关重要。
在CliniMACS设备上对9个干细胞移植物进行免疫磁珠纯化，并在TCRαβ/CD19清除前后进行流式细胞术质量控制（QC）。
作为一项挑战，建立了一种新的10色QC面板，可实现对移植物组成的准确量化。残留TCRαβ T细胞和CD19 B细胞的结合位点至少部分被清除抗体占据，从而妨碍了流式细胞术分析。基于各自的对照和假定的18%变异系数，残留TCRαβ T细胞的检测限为1细胞/微升，占CD45细胞的0.002%。TCRαβ和CD19细胞的对数清除率分别为-3.9和-3.3。干细胞、NK细胞和TCRγδ T细胞的回收率分别为82.1%、67.1%和72.7%。
在临床应用中，由于正确应用了所需的干细胞且T细胞剂量有限，该方法可能有助于改善移植结果。该面板专为TCRαβ/CD19清除后的QC设计，但也适用于其他清除策略。©2015国际临床细胞计量学会。
