Brun R, Jenni L, Schönenberger M, Schell K F
J Protozool. 1981 Nov;28(4):470-9. doi: 10.1111/j.1550-7408.1981.tb05322.x.
A series of new in vitro systems for the cultivation of bloodstream forms of Trypanosoma (Trypanozoon) brucei brucei, T. (T.) b. rhodesiense, and T. (T.) b. gambiense was developed. The standard system consists of a feeder layer of fibroblast-like cells derived from embryos of New Zealand White rabbits (REF( or a mountain vole, Microtus montanus (MEF), with HEPES-buffered Minimum Essential Medium (MEM), with Earle's salts, supplemented with 15% inactivated rabbit serum. These two and other feeder layers were cross-checked with different sera to test for growth support of bloodstream forms of the three trypanosome subspecies studied. Cultures could be initiated with bloodstream forms from mammalian hosts or from cryopreserved stabilates. Metacyclic forms from infected Glossina m. morsitans could also be sued as inoculum; they transformed within 6 h to bloodstream forms. Maintenance of cultures and growth properties are described in detail. Experiments were undertaken to confirm that the cultivated bloodstream forms still possess some fo the characteristic features of pleomorphic bloodstream populations. Cultivated bloodstream forms were always infective for mice, and a surface coat could be demonstrated by electron microscopy. They could also be cyclically transmitted through tsetse flies, and the metacyclic forms from these flies could be brought back into culture. In vitro cloning with single bloodstream forms and metacyclic forms could be achieve with high cloning efficiency. The consumption of glucose and the production of pyruvate and lactate were determined.
开发了一系列用于培养布氏锥虫（布氏锥虫属）、罗德西亚锥虫和冈比亚锥虫血流形式的新体外系统。标准系统由源自新西兰白兔胚胎的成纤维细胞样细胞饲养层（REF）或山地田鼠（MEF）组成，使用含有Earle's盐的HEPES缓冲的最低必需培养基（MEM），并补充15%的灭活兔血清。用不同血清对这两种及其他饲养层进行交叉检查，以测试对所研究的三种锥虫亚种血流形式的生长支持。培养物可以用来自哺乳动物宿主的血流形式或冷冻保存的稳定株起始。来自感染的采采蝇（采采蝇属）的循环后期形式也可用作接种物；它们在6小时内转化为血流形式。详细描述了培养物的维持和生长特性。进行实验以确认培养的血流形式仍具有多形性血流群体的一些特征。培养的血流形式对小鼠始终具有感染性，并且通过电子显微镜可以证明有表面被膜。它们也可以通过采采蝇进行循环传播，并且这些采采蝇的循环后期形式可以重新培养。用单个血流形式和循环后期形式进行体外克隆可以获得高克隆效率。测定了葡萄糖的消耗以及丙酮酸和乳酸的产生。
