Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Cytotherapy. 2009;11(7):923-35. doi: 10.3109/14653240903188921.
Clinical-scale lymphocyte enrichment from a leukapheresis product has been performed most routinely using costly magnetic bead separation systems that deplete monocytes, but this procedure may leave behind residual beads or antibodies in the enriched cell product. Counterflow centrifugal elutriation has been demonstrated previously to enrich monocytes efficiently for generation of dendritic cells. This study describes a modified elutriation procedure for efficient bead-free economical enrichment of lymphocytes from leukapheresis products from healthy donors and study subjects with human immunodeficiency virus (HIV) infection or malignancy.
Modified program settings and conditions for the CaridianBCT Elutra device were investigated to optimize lymphocyte enrichment and recovery. Lymphocyte enrichment was measured using a novel approach utilizing cell sizing analysis on a Beckman Coulter Multisizer and confirmed by flow cytometry phenotypic analysis.
Efficient enrichment and recovery of lymphocytes from leukapheresis cell products was achieved using modified elutriation settings for flow rate and fraction volume. Elutriation allowed for enrichment of larger numbers of lymphocytes compared with depletion of monocytes by bead adherence, with a trend toward increased lymphocyte purity and yield via elutriation, resulting in a substantial reduction in the cost of enrichment per cell. Importantly, significant lymphocyte enrichment could be accomplished using leukapheresis samples from healthy donors (n=12) or from study subjects with HIV infection (n=15) or malignancy (n=12).
Clinical-scale closed-system elutriation can be performed efficiently for the selective enrichment of lymphocytes for immunotherapy protocols. This represents an improvement in cost, yield and purity over current methods that require the addition of monocyte-depleting beads.
从白细胞分离产品中进行临床规模的淋巴细胞富集最常使用昂贵的磁珠分离系统来耗尽单核细胞，但该过程可能会在富集的细胞产物中留下残留的珠子或抗体。逆流离心洗脱已被证明可有效地富集单核细胞，用于生成树突状细胞。本研究描述了一种改良的洗脱程序，用于从健康供体和人类免疫缺陷病毒（HIV）感染或恶性肿瘤的研究对象的白细胞分离产品中高效、无珠且经济地富集淋巴细胞。
研究了 CaridianBCT Elutra 设备的改良程序设置和条件，以优化淋巴细胞的富集和回收。使用贝克曼库尔特 Multisizer 上的细胞大小分析的新方法测量淋巴细胞的富集，并通过流式细胞术表型分析进行确认。
使用改良的洗脱设置流速和分馏体积，可以从白细胞分离细胞产品中高效地富集和回收淋巴细胞。与通过珠附壁耗尽单核细胞相比，洗脱允许富集更多数量的淋巴细胞，并且通过洗脱，淋巴细胞纯度和产量呈增加趋势，导致每个细胞的富集成本显著降低。重要的是，通过使用健康供体（n=12）或 HIV 感染（n=15）或恶性肿瘤（n=12）的研究对象的白细胞分离样本，都可以实现显著的淋巴细胞富集。
临床规模的封闭系统洗脱可以高效地进行，用于免疫治疗方案中淋巴细胞的选择性富集。与需要添加耗尽单核细胞的珠子的现有方法相比，这在成本、产量和纯度方面都有所提高。
