Suen Y, Lee S M, Aono F, Hou S, Loudovaris M, Ofstein G, Bender J G
Nexell Therapeutics, Inc., Cellular Biology, 9 Parker, Irvine, CA 92618, USA.
Cytotherapy. 2001;3(5):365-75. doi: 10.1080/146532401753277184.
DC generated from monocytes have been used for vaccines. We have developed a monocyte enrichment procedure by depleting T and B cells with anti-CD2 and anti-CD19 Abs using the automated Isolex 300i magnetic cell selector for clinical-scale DC generation in gas permeable SteriCell culture bags. We have also compared DC function, yield and purity of DC generated from adherent monocytes using culture bags in a closed system, with DC generated in conventional tissue culture flasks.
Monocytes were enriched from normal donor apheresis products using CD2/19 depletion with experimental software on the Isolex 300i (ISO), adherence (AD) to SteriCell bags and to T175 flasks and then cultured for 7 days in serum-free X-VIVO 15 media with GM-CSF and IL-4. Phenotype and dextran uptake were analyzed by flow cytometry and allogeneic MLR was also evaluated.
ISO-DC and AD-DC from SteriCell bags showed similar viability. Higher purity of ISO-DC than AD-DC was measured by forward- and side-scatter flow cytometry. Similar expression of CD1a, CD80, CD86 and CD83 were observed in both ISO-DC and AD-DC. Similar dextran uptake and allo MLR were also observed.
These data indicated that functional DC were generated in gas permeable SteriCell culture bags from both ISO- and AD-monocytes in a closed system.
源自单核细胞的树突状细胞（DC）已被用于疫苗制备。我们开发了一种单核细胞富集程序，即使用自动化的Isolex 300i磁性细胞分选仪，通过用抗CD2和抗CD19抗体清除T细胞和B细胞，在透气的SteriCell培养袋中进行临床规模的DC生成。我们还比较了在封闭系统中使用培养袋从贴壁单核细胞生成的DC的功能、产量和纯度，与在传统组织培养瓶中生成的DC。
使用Isolex 300i（ISO）上的实验软件通过CD2/19清除法、使其贴壁于SteriCell袋和T175培养瓶，从正常供体的单采血液成分中富集单核细胞，然后在含GM-CSF和IL-4的无血清X-VIVO 15培养基中培养7天。通过流式细胞术分析表型和葡聚糖摄取情况，并评估异体混合淋巴细胞反应（MLR）。
来自SteriCell袋的ISO-DC和AD-DC显示出相似的活力。通过前向和侧向散射流式细胞术测量，ISO-DC的纯度高于AD-DC。在ISO-DC和AD-DC中均观察到CD1a、CD80、CD86和CD83的相似表达。也观察到相似的葡聚糖摄取和异体MLR。
这些数据表明，在封闭系统中，功能性DC可从透气的SteriCell培养袋中的ISO-单核细胞和AD-单核细胞生成。
