摘要
通过比较溶液的离子浓度对病毒形态结构的影响，发现GCHV在低盐溶液低温（-20℃）保存条件下，外壳蛋白会自动降解，但不十分完全。采用含1％NP40（Nonidetp40）的稀盐溶液处理感染病毒的细胞，经冻融及差异离心，电镜下观察到纯度较高的病毒双层衣壳、核心衣壳及散在的子粒。纯化的病毒亚单位用核酸酶消化后，与等量的福氏佐剂混合，免疫家兔制备抗体。该抗血清经中和试验测定，显示了较强的中和病毒的能力。纯化的病毒亚单位在7％聚丙烯酰胺凝胶电泳条件下，未见病毒电泳条带。同时采用地高辛标记的GCHVcDNA克隆探针，与病毒RNA及亚单位样品进行斑点杂交，检测病毒RNA灵敏度达到10pg．病毒亚单位样品均为阴性结果。该亚单位制剂具有纯度高、体积小的特点，将是一种安全性能好、使用方法简便、又便于保存和运输的亚单位疫苗。
By studying the flue structure of GCHV (Grass Carp Hemorrhage Virus), It was found that the capsid of purified GCHV can be degraded spontaneously into its morphological units by prologing in a buffer of low ionic strength, but not very completely. The virus preparation was obtained from GCHV infected cell, then treated with 1% NP40 reagent in low-salt solution and fieezed thaw ng, finally purified by differential centrifugation. Using negative staining method, out, core capsid and capsomeres of the virus were showed by electron micrograph. Afterwards, the preparation digested by RNase was equally mixed with Freund's adjuvant and immunized rabbits. The antibody was tested by ELISA and neutralization test, which demostrated the preparation can elicit good levels of antibody. The RNA-free preparation was detected by PAGE and DIG-labelled GCHV-cDNA hybridization. The sensitivity of the test is approximately to 10pg, the samples of subunit all showed negative. The preparation has a high purity, small volume and better safety. Besides, it is easy to use, also convenient in transportation and conservation. It will be a ideal vaccine against grass carp hemorrhage disease.
出处
《中国病毒学》
CSCD
1997年第1期77-82,共6页
Virologica Sinica
