猪繁殖与呼吸综合征病毒单克隆抗体的制备及鉴定

Development and Identification of Monoclonal Antibodies Against Porcine Reproductive and Respiratory Syndrome Virus

将猪繁殖与呼吸综合征病毒(PRRSV)经密度梯度离心纯化后免疫Balb/c小鼠,取免疫小鼠脾细胞与SP2/0细胞融合,间接ELISA筛选能稳定分泌抗PRRSV单克隆抗体的杂交瘤细胞株,并对制备出的单克隆抗体进行生物学与免疫学特性鉴定。结果成功筛选出3株能分泌IgG型单克隆抗体的杂交瘤细胞株,分别命名为6A4、3F1和7F2。3株抗体均表现良好的特异性,ELISA方法检测与其他病毒及Marc-145细胞培养物无交叉反应,免疫荧光染色试验表明3株单克隆抗体均可不同程度地识别细胞中的病毒而呈现特异性荧光。Western blot说明6A4、7F2能够分别特异性识别25ku和15ku蛋白。试验初步表明6A4、3F1具有中和抗体活性,7F2不具有抗体依赖性增强(antibody dependent enhancement,ADE)作用,但可以应用于PRRSV N蛋白检测。

Monoclonal antibodies （mAb ） against porcine reproductive and respiratory syndrome virus （PRRSV） were developed from the mice immunized with the virus purified by differential and sucrose den-sity gradient centrifugations .Three mAb were prepared and characterized ,named 6A4 ,3F1 and 7F2 .All of the mAb showed good specificity ,which could specifically react to PRRSV but not cross react to other pig viruses . Perinuclear fluorescence was found in varying degrees by immunofluorescence staining , showed that all the three mAb can recognize PRRSV in infected cells .Among the 3 mAb in a Western blot assay ,6A4 recognized a 25 ku band of PRRSV and 7F2 recognized 15 ku .By the ADE and antibody neu-tralization tests ,6A4 and 3F1 showed neutralization activity ,and 7F2 showed no ADE activity but poten-tial application in N protein detection by ELISA .