[摘 要] 目的：比较国产与进口CD3/CD28激活磁珠制备的CAR-T细胞的性能差异，为国内CAR-T细胞研发和生产提供备用 或替代方案。方法：以进口CD3/CD28激活磁珠︰CD3+ T细胞比例为1∶1的成熟方案作为研究对照，国产磁珠设置1∶2、1∶1 和2∶1的比例梯度激活T细胞，T细胞激活72 h后使用CAR慢病毒感染制备CAR-T细胞。分别在慢病毒感染后的第2、4、7天计 数，监测CAR-T细胞增殖情况；感染5 d后采用流式细胞术检测CAR-T细胞阳性率；感染8 d后采用流式细胞术检测CAR-T细胞 的CD4/CD8分型情况和PD1+ TIM3+耗竭细胞比例。结果：国产CD3/CD28激活磁珠与进口磁珠制备的CAR-T细胞CD4/CD8分 型相似，CAR-T细胞阳性率略低于进口磁珠（53.7% vs 57.9%），但CAR-T细胞扩增较进口磁珠增加1倍左右，耗竭水平仅为进口 磁珠的一半（4.21% vs 7.91%）。此外，国产磁珠的使用量低于进口磁珠，有利于降低CAR-T细胞研发和生产成本。结论：国产 CD3/CD28激活磁珠用于制备CAR-T细胞总体性能不劣于进口磁珠，有潜力作为进口磁珠的备用或替代方案。

[Abstract] Objective: To compare the performance differences of domestic and imported CD3/CD28 activation beads for manufacturing CAR-T cells, providing a backup or alternative for domestic CAR-T cell research and manufacture. Methods: A mature protocol using imported CD3/CD28 activation beads with a 1∶1 ratio for CD3+ T cells was implemented as research control. Domestic beads were used with gradient ratios of 1∶2, 1∶1, and 2∶1 to activate T cells. 72 h after T cell activation, CAR-T cells were manufactured by CAR lentiviral infection and cell proliferation was monitored at 2-, 4-, and 7-days post-infection. Flow Cytometry was used to detect CAR-T cell positivity 5 days after infection and to detectCD4/CD8 phenotype of CAR-T cells and PD1+ TIM3+ cell exhaustion ratio 8 days after infection. Results: CAR-T cells manufactured by domestic CD3/CD28 activation beads exhibited similar phenotype compared with those manufactured by imported CD4/CD8 beads. The positive rate of CAR-T cells prepared with domestic beads was slightly lower than that of imported beads (53.7% vs 57.9%). However, the proliferation of CAR-T cells manufactured by domestic beads was about twice that of cells manufactured by imported beads, and the exhaustion level was only half that of imported beads (4.21% vs 7.91%). Moreover, the use of domestic magnetic beads was lower than that of imported magnetic beads, which was advantageous for cutting the costs of CAR-T cells research and manufacture. Conclusion: Domestic CD3/CD28 activation beads used for CAR-T cells manufacturing demonstrate comparable overall performance to their imported counterparts, showing potential as a backup or alternative for imported beads.

国家自然科学基金（No. 82304535）