A comparative analysis of Freon substitutes in the purification of reovirus and calicivirus
A comparative analysis of Freon substitutes in the purification of reovirus and calicivirus
复制标题
DOI：
10.1016/s0166-0934(00)00217-2
复制
发表时间：
2000-10-01
影响因子：
3.1
通讯作者：
Coombs, KM
中科院分区：
文献类型：
--
作者：
Mendez, II;Hermann, LL;Coombs, KM
氟利昂113（氟利昂）是一种重要的成分，用于一些病毒纯化方法，以分离病毒从感染的细胞碎片。由于其环境和毒性危害，氟利昂的可用性有限，并且受到更严格的监管。选择几种有机溶剂替代物以鉴定用于纯化可培养呼肠孤病毒和挑剔杯状病毒的合适的氟利昂替代物。用测试的每种溶剂从组织培养的细胞中提取呼肠孤病毒，并通过标准技术在氯化铯梯度中纯化。通过形态学检查和感染性研究分析纯化的病毒粒子的物理和生物学特性的保守性。还检查了使用所选溶剂从粪便样品中纯化杯状病毒核酸。将溶剂提取的杯状病毒RNA逆转录，并通过标准诊断性117 bp扩增子的聚合酶链反应扩增进行定量。这些研究表明，Vertrel（R）XF（一种新开发的环保型氟利昂替代品）和异戊烷/1-氯丁烷的7：3混合物是合适的替代品。考虑到易燃性和易用性，Vertrel（R）XF是纯化这些无包膜病毒的首选氟利昂替代品。(C)2000 Elsevier Science B.V.保留所有权利。
Freon 113 (Freon) is an essential component used in some viral purification methods to separate virus from infected cell debris. With its environmental and toxic hazards, Freon's availability is limited and more tightly regulated. Several organic solvent substitutes were selected to identify a suitable Freon replacement for the purification of both cultivable reovirus and fastidious calicivirus. Reovirus was extracted from tissue cultured cells with each solvent tested and purified in cesium chloride gradients by standard techniques. Purified virions were analyzed for conservation of physical and biological properties by morphological examination and infectivity studies. The purification of calicivirus nucleic acid from stool samples using selected solvents was also examined. Solvent-extracted calicivirus RNA was reverse transcribed and quantified by polymerase chain reaction amplification of a standard diagnostic 117 bp amplicon. These studies indicated that Vertrel(R) XF (a newly developed environmentally friendly Freon substitute) and a 7:3 mixture of isopentane/1-chlorobutane are suitable replacements. Considerations of flammability and ease of use suggest that Vertrel(R) XF is the preferred choice as a Freon substitute for the purification of these non-enveloped viruses. (C) 2000 Elsevier Science B.V. All rights reserved.