摘要
【目的】针对O型口蹄疫病毒(Foot-and-mouth Disease Virus,FMDV)制备特异性的单克隆抗体,为进一步研制O型口蹄疫诊断方法提供物质基础。【方法】用纯化的O型口蹄疫病毒免疫Balb/c小鼠,取脾细胞与SP2/0骨髓瘤细胞进行融合,结合间接ELISA和间接免疫荧光(IFA)筛选,有限稀释法克隆,建立稳定的阳性杂交瘤细胞株,并对制备的单克隆抗体进行生物学特性鉴定。【结果】获得了2株稳定分泌单克隆抗体的杂交瘤细胞株,分别命名为2B9和5F7,抗体亚类鉴定其均为IgG1/κ类型;IFA结果显示,2株单克隆抗体仅与O型FMDV反应,不与Asia1型FMDV反应,推测其均为抗O型FMDV的型特异性单克隆抗体;Western blot结果显示,2株单克隆抗体均无特异性条带出现,表明其所针对的抗原表位均为构象型表位;相加ELISA试验表明,2B9与5F7两株单克隆抗体识别的抗原表位相近或相同;经硫氰酸盐洗脱法测定,2B9和5F7的相对亲和力指数均为1.5 mol/L;中和试验显示,2株单克隆抗体都不具有中和活性。【结论】2株单克隆抗体的获得及其生物学性质的鉴定,为O型口蹄疫诊断方法的建立提供了基础材料。
[Objective] Specific monoclonal antibodies （mAbs） against FMDV O was prepared so as to provide materials for the study of diagnostic method of FMD. [Method] By fusing SP2/0 myeloma cells and spleen cells of Balb/c mice immunized with purified virus preparations with the standard hybridomas technique,indirect ELISA, indirect immunofluorescence assay（IFA） and limited dilution for cell cloning were performed to screen positive hybridomas,then the biological characteristics of the mAbs were identi- fied. [Result] Two stable hybridoma cell strains, named 2B9 and 5F7, were established. These mAbs were IgG1/k isotype and contained kappa light chains. IFA showed that they reacted with FMDV serotype O,but not with FMDV serotype Asial. Western blot analysis suggested that the epitopes they recognized were conformational since any specific band didn＇t appear. The additive ELISA showed that antigen epitopes recognized by 2B9 and 5F7 could be close to each other. Their relative affinity index were 1.5 mol/L,which was indicated by thiocyanate elution measurement. Neutralization test showed that they didn＇t show any neutralizing activity to O serotype FMDV strains. [Conclusion] These two mAbs could be used as experimental materials for the establishment of the diagnostic method of FMD.
出处
《西北农林科技大学学报（自然科学版）》
CSCD
北大核心
2008年第11期35-39,46,共6页
Journal of Northwest A&F University(Natural Science Edition)
基金
黑龙江省"十一五"重大项目(GA06B202)
关键词
O型口蹄疫病毒
单克隆抗体
生物学特性鉴定
foot-and-mouth disease virus serotype O
monoclonal antibody
identification of bilolgical characteristic
