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Cell separation improves the sensitivity of detecting rare human normal and leukemic hematopoietic cells in vivo in NOD/SCID mice
Cytotherapy ( IF 4.5 ) Pub Date : 2000-06-01 , DOI: 10.1080/146532400539350
T L Holyoake 1 , C Horrocks , T Thomas , C J Eaves , A C Eaves
Affiliation  

BACKGROUND This report describes a novel cell-separation procedure developed to improve detection and analysis of rare human hematopoietic populations, obtained from NOD/SCID mice engrafted with normal and/or leukemic stem cells. METHODS In preliminary experiments, artificial mixtures of murine and human BM cells were labeled with a combination of Abs specific for murine hematopoietic cells, prior to immunomagnetic negative selection using StemSep. In subsequent experiments, BM was harvested from individual NOD/SCID mice transplanted 6-12 weeks earlier with either human cord blood or primary CML cells and a similar immunomagnetic selection procedure was applied to enrich human cells present. RESULTS Application of this selection procedure to mixtures of murine and human hematopoietic cells using anti-mouse CD45 and Ter-119 allowed a > 1000-fold depletion of murine cells with > 50% recovery of human cells, including progenitors. This level of depletion and recovery were found to be reproducible for NOD/SCID mice transplanted and engrafted with human cord blood stem cells, thus facilitating detection of human progenitors, including colony-forming cells (CFC) and LTCIC. For NOD/SCID mice previously transplanted with CML cells, this procedure increased the sensitivity of detecting rare human cell subsets by up to > 100-fold. This, in turn, improved the sensitivity of RT-PCR for BCR-ABL and made possible the identification by FACS of various minor subsets of human cells, including CD34(-)CD19/20(+) B-lineage cells, CD34(+) progenitors, mature CD15(+) myeloid cells and CD3(+) T cells present in the mice. DISCUSSION This simple cell-depletion procedure should facilitate future investigations of normal and CML stem cell populations in vitro and in NOD/SCID mice.

中文翻译:

细胞分离提高了在 NOD/SCID 小鼠体内检测稀有人类正常和白血病造血细胞的灵敏度

背景本报告描述了一种新的细胞分离程序,该程序旨在改进对从移植有正常和/或白血病干细胞的 NOD/SCID 小鼠中获得的稀有人类造血细胞群的检测和分析。方法 在初步实验中,在使用 StemSep 进行免疫磁性负选择之前,用小鼠造血细胞特异性抗体组合标记小鼠和人类 BM 细胞的人工混合物。在随后的实验中,从 6-12 周前用人脐带血或原代 CML 细胞移植的个体 NOD/SCID 小鼠中收获了 BM,并应用了类似的免疫磁性选择程序来丰富存在的人类细胞。结果 将此选择程序应用于使用抗小鼠 CD45 和 Ter-119 的鼠和人造血细胞混合物允许 > 鼠细胞耗竭 1000 倍,而人类细胞(包括祖细胞)的恢复率超过 50%。发现这种消耗和恢复水平对于移植和植入人类脐带血干细胞的 NOD/SCID 小鼠是可重现的,从而促进人类祖细胞的检测,包括集落形成细胞 (CFC) 和 LTCIC。对于先前移植了 CML 细胞的 NOD/SCID 小鼠,此过程将检测稀有人类细胞亚群的灵敏度提高了 100 倍以上。这反过来又提高了 RT-PCR 对 BCR-ABL 的敏感性,并使通过 FACS 鉴定各种次要的人类细胞亚群成为可能,包括 CD34(-)CD19/20(+) B 谱系细胞、CD34(+ ) 祖细胞、成熟的 CD15(+) 骨髓细胞和 CD3(+) T 细胞存在于小鼠中。
更新日期:2000-06-01
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