猪O型口蹄疫病毒非结构蛋白3ABC抗原模拟表位的筛选

Screening of Mimotopes for Non-structural Protein 3ABC of O Type Foot-and-mouth-disease Virus

口蹄疫病毒(FMDV)非结构蛋白(NSP)3ABC与FMDV复制有关,感染FMDV的动物产生的NSP 3ABC抗体可在体内存留较长时间,是鉴别诊断动物接种疫苗与自然感染口蹄疫的可靠指标。本文从猪FMDV-NSP3ABC阳性抗血清中分离和纯化IgG,以此为固相筛选分子,对噬菌体随机十二肽库进行4轮吸附-洗脱-扩增的富集筛选后,随机挑取20个噬菌斑进行扩增,用ELISA方法分别检测扩增后的噬菌体抗原性,其中有8个噬菌体克隆与纯化的IgG有较强的特异性结合能力;对得到的阳性克隆提取ssDNA进行测序,分析所递呈的氨基酸序列,其中的7个噬菌体展示肽的氨基酸片段具有较高的保守性;进一步分别以8个阳性噬菌体克隆为固相捕获分子,对22份疑似FMD病猪血清进行检测,结果显示,有5个噬菌体克隆检测结果与试剂盒检测有较高的符合率。本研究为FMDV-NSP 3ABC抗原表位结构进一步研究和建立猪自然感染FMDV快速鉴别诊断新方法奠定了基础。

In this article, the IgG was purified from the positive antiserum to FMDV-NSP 3ABC. The IgG was used as target molecule to screen the phage display 12-met random peptide library for 4 rounds by the hinding-eluting-amplification method. After 4 rounds of screening, 20 clones were picked out randomly and amplified. Their antigenicities were tested by ELISA. Among 20 phage clones, 8 phage clones had specific reactions with the purified IgG. The ssDNA of the positive clones were abstracted for DNA sequencing and the sequences of peptides displayed on the positive clones were analyzed by ExPASy method. The results showed that the peptides displayed on the 7 phage clones were highly homologous. 22 duhitahle porcine sera were tested by 8 positive clones which were used as target molecules respectively, and 5 clones showed high correlation with traditional Kit. These results were successfully provided a foundation for research of the structural epitope of the FMDV-NSP 3ABC and to establish a new diagnostic method for the natural infection of the FMDV.