Kepley C, Craig S, Schwartz L
Department of Microbiology and Immunology, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298.
J Immunol Methods. 1994 Sep 30;175(1):1-9. doi: 10.1016/0022-1759(94)90326-3.
Basophils typically account for approximately 1% of the white cells in peripheral blood. We have developed a unique method for purifying basophils from whole blood of normal subjects to at least 95% purity. Basophils are separated from other cell types by density-dependent sedimentation in Percoll and cell sorting, based solely on their size and granularity. The mean overall yield ranged from 5% to 28%. The procedure is typically completed within 4 h. The highly purified basophils obtained are functionally competent and morphologically intact. They release histamine in response to Fc epsilon RI-mediated stimulation, express Fc epsilon RI and BSP-1 ligand as analyzed by flow cytometry, and exhibit the known characteristic ultrastructural features of basophils by electron microscopy. This procedure avoids positive-selection antibodies that might perturb receptors on basophils or negative-selection antibodies that might activate other cell types, and can be used to obtain basophils for studies in vitro.
嗜碱性粒细胞通常占外周血白细胞的约1%。我们开发了一种独特的方法，可从正常受试者的全血中纯化嗜碱性粒细胞，纯度至少达到95%。嗜碱性粒细胞通过在Percoll中基于密度的沉降和细胞分选与其他细胞类型分离，仅基于它们的大小和颗粒度。平均总回收率为5%至28%。该过程通常在4小时内完成。获得的高度纯化的嗜碱性粒细胞功能正常且形态完整。它们在FcεRI介导的刺激下释放组胺，通过流式细胞术分析表达FcεRI和BSP-1配体，并通过电子显微镜呈现嗜碱性粒细胞已知的特征性超微结构特征。该方法避免了可能干扰嗜碱性粒细胞上受体的阳性选择抗体或可能激活其他细胞类型的阴性选择抗体，可用于获取嗜碱性粒细胞进行体外研究。
