Graziani-Bowering G M, Graham J M, Filion L G
Department of Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ont., Canada.
J Immunol Methods. 1997 Sep 24;207(2):157-68. doi: 10.1016/s0022-1759(97)00114-2.
A commercial monocyte isolation technique based on the OptiPrep density-gradient medium was up-scaled with respect to sample and reagent volumes. The results of 7 isolations are reported in which the average purity ranged from 87.9 to 96.4%. In all but the initial isolation, monocytes were defined as CD15+ dim CD4+ dim as assessed by flow cytometric analysis; in the first isolation, monocytes were defined by the traditional CD14+ CD4+ dim combination. The mean yield (the number of isolated monocytes relative to the number present in the buffy coat) of all isolations was 26.1%, with the individual yields ranging from 10.8 to 41.4%. The mean number of isolated monocytes per experiment was 3.6 x 10(6) monocytes for those isolations performed using 14 ml of buffy coat/OptiPrep mixture (n = 4). The isolated cells were viable (> 95%) and were not activated, according to HLA-DR expression. This technique is a convenient, tast (less than 2 h), relatively simple, and inexpensive alternative to traditional monocyte isolation techniques. The up-scaled version of this method also results in significantly higher numbers of monocytes per isolation than some traditional techniques. Furthermore, this is the first literature report of the use of the OptiPrep density-gradient medium for the isolation of monocytes.
一种基于OptiPrep密度梯度介质的商业化单核细胞分离技术在样本和试剂体积方面进行了扩大。报告了7次分离的结果，其中平均纯度范围为87.9%至96.4%。除了首次分离外，通过流式细胞术分析，单核细胞被定义为CD15+ 弱阳性CD4+ 弱阳性；在首次分离中，单核细胞由传统的CD14+ CD4+ 弱阳性组合定义。所有分离的平均产量（分离的单核细胞数量相对于血沉棕黄层中存在的数量）为26.1%，个体产量范围为10.8%至41.4%。对于使用14 ml血沉棕黄层/OptiPrep混合物进行的分离实验（n = 4），每次实验分离的单核细胞平均数量为3.6×10⁶个单核细胞。根据HLA-DR表达，分离的细胞具有活力（> 95%）且未被激活。该技术是一种方便、耗时短（少于2小时）、相对简单且廉价的传统单核细胞分离技术的替代方法。该方法的扩大版本每次分离得到的单核细胞数量也比一些传统技术显著更多。此外，这是关于使用OptiPrep密度梯度介质分离单核细胞的首次文献报道。
