猪繁殖与呼吸综合征病毒凝胶层析纯化方法的建立与初步应用  

Establishment and Preliminary Application of Gel Chromatography Purification Method for Porcine Reproductive and Respiratory Syndrome Virus

作　　者：孙悦 王尚辉 涂亚斌[1] 杨勇博 王洪峰 夏大松 蔡雪辉[1] 安同庆[1] 孙明霞 SUN Yue;WANG Shang-hui;TU Ya-bin;YANG Yong-bo;WANG Hong-feng;XIA Da-song;CAI Xue-hui;AN Tong-qing;SUN Ming-xia(State Key Laboratory of Veterinary Biotechnology,Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin 150069,China;Harbin Weike Biotechnology Co.,Ltd.,Harbin 150069,China)

机构地区：[1]中国农业科学院哈尔滨兽医研究所、兽医生物技术国家重点实验室,黑龙江哈尔滨150069 [2]哈尔滨维科生物技术有限公司,黑龙江哈尔滨150069

出　　处：《中国兽医杂志》2023年第4期1-6,共6页Chinese Journal of Veterinary Medicine

基　　金：国家自然科学基金(32072851);兽医生物技术国家重点实验室开放课题(SKLVBF202110)。

摘　　要：为了高效分离纯化并获得结构完整的猪繁殖与呼吸综合征病毒(PRRSV),用于PRRSV的形态学研究、动物免疫以及制备高质量疫苗,本试验在Marc-145细胞中增殖PRRSV HuN4株,经超滤浓缩后,分别采用氯化铯密度梯度离心方法和凝胶层析技术纯化PRRSV。结果显示,与氯化铯密度梯度离心相比,凝胶层析纯化可以得到纯度更高、结构更完整的病毒粒子。将凝胶层析纯化得到的PRRSV免疫小鼠制备多抗,间接免疫荧光试验(IFA)结果显示,多抗血清可与PRRSV发生特异性反应;酶联免疫吸附试验(ELISA)结果显示,抗体效价可达到1∶25600以上,纯化的病毒具有良好的免疫原性。结果表明,应用凝胶层析技术可得到高纯度、结构完整的PRRSV粒子,且纯化后PRRSV能够诱导良好的免疫应答,本试验结果可为PRRSV的纯化提供参考。In order to isolate,purify and obtain structurally complete porcine reproductive respiratory syndrome virus(PRRSV)for morphological study,animal immunization and preparation of high-quality vaccine,PRRSV HuN4 strain was proliferated on Marc-145 cells.After concentration by ultrafiltration,PRRSV was purified by cesium chloride gradient centrifugation and gel chromatography.The results showed that higher purity and more intact virion could be obtained by gel chromatography than by cesium chloride density gradient centrifugation.Mice were immunized with gel chromatography-purified PRRSV for polyclonal antibody production.Indirect immunofluorescence assay(IFA)showed that the resulting polyclonal antibodies reacted with PRRSV specifically,and enzyme linked immunosorbent assay(ELISA)showed the serum antibody titers reached more than 1∶25600,confirming the excellent immunogenicity of the purified virus.Thus,PRRSV with high purity,complete structure is obtainable by gel chromatography for induction of strong immune response.The results of this test provide an important reference for PRRSV purification.

关 键 词：猪繁殖与呼吸综合征病毒 凝胶层析技术 纯化 形态学 

分 类 号：S855.3[农业科学—临床兽医学]