Dooley D C, Simpson J F, Meryman H T
Exp Hematol. 1982 Aug;10(7):591-9.
Techniques were developed for the isolation of human neutrophils from large volumes of human blood. This preparative procedure avoids exposure to Ficoll/Hypaque and offers a low cost alternative to centrifugal elutriation. First, leukocyte-enriched suspensions were obtained by treating buffy coats with 2% dextran. The leukocyte suspensions were then fractionated on three steps Percoll density gradients. The gradients were designed to carry large numbers of cells while maintaining resolution of the cell peaks. As a result, at least 320 X 10(6) leukocytes could be processed on a single 12 ml gradient. Extensive testing of the procedure showed that the purity of the neutrophil band was 96.5 +/- 3.16% (n = 50). The band contained 82.4 +/- 14.6% (n = 39) of the neutrophils applied to the gradient. Viability assays (bacterial killing, chemotaxis) demonstrated that neutrophils isolated on Percoll gradients were just as active as those harvested by centrifugal elutriation. Recovery data are presented and compared to the results obtained by others.
已开发出从大量人血中分离人中性粒细胞的技术。该制备方法避免了接触Ficoll/Hypaque，为离心淘洗提供了一种低成本替代方案。首先，通过用2%葡聚糖处理血沉棕黄层获得富含白细胞的悬浮液。然后将白细胞悬浮液在三步Percoll密度梯度上进行分级分离。这些梯度旨在承载大量细胞，同时保持细胞峰的分辨率。结果，在单个12 ml梯度上至少可处理320×10⁶个白细胞。对该方法的广泛测试表明，中性粒细胞带的纯度为96.5±3.16%（n = 50）。该带包含应用于梯度的中性粒细胞的82.4±14.6%（n = 39）。活力测定（细菌杀伤、趋化性）表明，在Percoll梯度上分离的中性粒细胞与通过离心淘洗收获的中性粒细胞一样活跃。给出了回收率数据并与其他人获得的结果进行了比较。
