Vaisman B, Konijn A M, Fibach E
Department of Hematology, Hadassah University Hospital, Faculty of Medicine, Jerusalem, Israel.
Acta Haematol. 1999;101(3):135-9. doi: 10.1159/000040939.
The two-phase liquid culture, which supports the development of human erythroid progenitors into hemoglobin-containing orthochromatic normoblasts, provides high yield of erythroblasts with synchronized development. However, other cell types are also present in the culture. In the present report, we used immunomagnetic separation for the purification of the developing erythroid precursors. At different stages of the culture, aliquots of the cells were incubated with anti-glycophorin A, B monoclonal antibodies, followed by anti-mouse IgG-coated magnetic beads, and separated by a magnet. In mature cultures, all isolated glycophorin-positive cells contained hemoglobin, as indicated by their staining with benzidine. All glycophorin-negative cells were benzidine negative. In earlier cultures, morphological examination and measurements of hemoglobin content showed that the erythroid precursors could be isolated at different stages of maturation. Thus, the combination of the liquid culture procedure with the immunomagnetic separation technique permitted to obtain large samples of pure erythroid cells, which were synchronized at subsequent stages of maturation. The method enables comprehensive studies of developing erythroid cells from normal donors as well as from patients with various disorders of erythropoiesis.
两阶段液体培养可支持人类红系祖细胞发育为含血红蛋白的正染性晚幼红细胞，能高产同步发育的成红细胞。然而，培养物中也存在其他细胞类型。在本报告中，我们使用免疫磁珠分离法纯化发育中的红系前体细胞。在培养的不同阶段，将细胞等分试样与抗血型糖蛋白A、B单克隆抗体孵育，然后与抗小鼠IgG包被的磁珠孵育，并通过磁铁进行分离。在成熟培养物中，所有分离出的血型糖蛋白阳性细胞都含有血红蛋白，这通过它们用联苯胺染色得以表明。所有血型糖蛋白阴性细胞联苯胺染色均为阴性。在早期培养物中，形态学检查和血红蛋白含量测定表明，红系前体细胞可在不同成熟阶段被分离出来。因此，液体培养程序与免疫磁珠分离技术的结合能够获得大量处于后续成熟阶段同步化的纯红系细胞样本。该方法能够对正常供体以及患有各种红细胞生成障碍疾病患者的发育中的红系细胞进行全面研究。
