鉴别狂犬病病毒强弱毒株中和性单克隆抗体的制备及初步应用

为获得对狂犬病病毒弱毒疫苗株筛选和糖蛋白抗原结构分析的单克隆抗体，将鹿狂犬病病毒 ８２０２ 株在适宜的条件下培养 ７２ ｈ，收集无细胞上清，经离心沉淀、 Ｚｎ （ Ａ Ｃ）２ 浓缩、１０％ ～５０％ 质量浓度的蔗糖密度梯度离心纯化、 Ｓ Ｄ Ｓ Ｐ Ａ Ｇ Ｅ 分析证明，产物中富含狂犬病病毒特异的结构蛋白。以上述纯化病毒免疫 Ｂ Ａ Ｌ Ｂ／ｃ 小鼠的脾细胞与 Ｓ Ｐ２／０ 骨髓瘤细胞融合，经 ３～５ 次克隆，间接 Ｅ Ｌ Ｉ Ｓ Ａ 筛选， Ｗ ｅｓｔｅｒｎ ｂｌｏｔｔｉｎｇ 鉴定，并与不同科病毒反应，建立了 ２ 株（４ Ａ５ ，３ Ａ５ ）可稳定分泌抗狂犬病病毒 Ｇ 蛋白的杂交瘤细胞株。这 ２ 株 Ｍ ｃ Ａｂ 与 ４株狂犬病病毒反应，经间接 Ｅ Ｌ Ｉ Ｓ Ａ 检测，发现能与强毒株反应，而与弱毒株不反应。细胞中和试验证实，这 ２株 Ｍ ｃ Ａｂ 具有良好的细胞中和活力。

Preparation and Primary Application of Neutralizing Monoclonal Antibodies for Distinguishing Attenuated Rabies Viruses from Virulent Ones

Rabies virus cell adapted strain 8202 (isolated from sika deer) was cultured in suitable condition and purified by Zn(AC) 2 precipitation and subsequently sucrose gradient centrifugation. Two hybridomas cell strains secreting specific McAbs against rabies virus glycoprotein were obtained and were characterized by Western blotting assay. The specificity of these McAbs were identified using 4 rabies virus strains (an attenuated vaccine strain SRV 9, CVS, 8202 brain strain and 8202 cell strain), and showed remarkable difference between them. All McAbs could react with CVS, 8202 cell strain and 8202 brain strain, however neither of them showed reactivity with SRV 9. Highly cellular neutralizing abilities were also demonstrated on them. The 2 anti glycoprotein McAbs showing different reactivity characteristics among 4 rabies virus strains could be used to distinguish attenuated strains from virulent ones.