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制备抗Ⅱ型单纯疱疹病毒(Simplex Herpes VirusⅡ,HSV-2)表面抗原gD的单克隆抗体(mAb)并对其性质进行鉴定。以基因工程重组制备纯化的HSV2-gD为抗原免疫BALB/c小鼠,采用常规融合制备杂交瘤细胞,通过HAT选择培养、有限稀释法获得单克隆细胞株,用间接ELISA法、Western blot方法筛选和鉴定阳性杂交瘤细胞株。获得了4株可稳定分泌抗HSV2-gD抗体的杂交瘤细胞株,分别命名为1A11C6、1A11F10、4C9D5以及4C9E9,抗体的类型均为IgG1,轻链均为κ型,Western blot结果显示各单抗都可以特异性识别HSV2-gD。用杂交瘤细胞注射小鼠制备腹水,腹水内的抗体效价均大于1×10-5,腹水内的抗体经硫酸铵-辛酸分级沉淀和阴离子交换柱纯化,成功制备了纯度超过95%的高特异性抗体,抗体有较好的特异性。为进一步研究HSV2感染和临床上的预防、诊断及治疗提供了有力的工具。
Monoclonal antibodies (mAbs) against the surface antigen gD of Simplex Herpes Virus II (HSV-2) were prepared and their properties were identified. BALB / c mice were immunized with recombinant HSV2-gD recombinant protein by genetic engineering, and hybridoma cells were prepared by conventional fusion. HAT cells were selected for culture. Monoclonal cell strains were obtained by limiting dilution method. Indirect ELISA and Western blot were used to screen And identification of positive hybridoma cell lines. Four hybridoma cell lines stably secreting anti-HSV2-gD antibody were obtained and designated as 1A11C6, 1A11F10, 4C9D5 and 4C9E9, respectively. The types of antibodies were IgG1 and the light chains were both κ type. Western blot showed that each monoclonal antibody HSV2-gD can be specifically recognized. The ascites was prepared by injecting mice with hybridoma cells. The antibody titers in ascites were all more than 1 × 10-5. The antibodies in ascites were purified by ammonium sulfate-octanoic acid fractionation and anion exchange chromatography. The purity of over 95% Specific antibodies, antibodies have better specificity. It provides a powerful tool for further study of HSV2 infection and clinical prevention, diagnosis and treatment.