人乳头瘤病毒(HPV)31型中和单克隆抗体的制备及性质鉴定

目的:构建获得HPV31构象依赖的中和单抗。方法:采用昆虫细胞表达的HPV31 L1VLP(virus-like particle, VLP)免疫BALB/c小鼠,取免疫鼠脾细胞与SP2/0细胞融合,收集杂交瘤细胞培养上清,首先采用HPV31 VLP-ELISA及HPV31假病毒中和实验筛选分泌HPV31中和单抗的杂交瘤细胞株,然后纯化单克隆抗体,分别采用假病毒中和实验及ELISA实验对纯化后单克隆抗体进行鉴定,包括抗体的亚型及其结合表位的构象特征、针对HPV31的中和IC50及针对HPV16、HPV18、HPV33、HPV45、HPV52、HPV58、HPV6、HPV11的交叉中和活性。结果:筛选获得9株HPV31构象依赖中和单抗,其中3株HPV31特异性中和单抗中有2株的IC5010 ng/mL,分别是XM31-13(0.36,Ig G1)及XM31-23(7.10,Ig G1),6株交叉中和单抗中亦有2株的IC5010ng/mL,分别是XM31-19(7.14,Ig G1)、XM31-20(6.91,Ig G1)。结论:获得的9株HPV31构象依赖的中和单抗,特别是其中4株单抗IC50值10 ng/mL的4株单抗,可用于含HPV31L1VLP多价疫苗的质控疫苗的研究。

Preparation and Characterization of Neutralizing Monoclonal Antibodies Against Human Papillomavirus Type 31

ABSTRACT Objective: Preparation of some conformation-dependent neutralizing monoclonal antibodies (mAbs) against HPV31. Methods: Female BALB/c mice were immunized with HPV31 L1 virus-like particles (VLPs) expressed by insect cells, and spleenocytes of the zimmunized mice were fused with SP2/0 cells to prepare hybridoma cells, the culture supernatants were-collected for identification of neutralizing mAb secreting cells by HPV31 VLP-ELISA and HPV31 pseudovirus neutralization experiments. Monoclonal antibodies were purified and identified by ELISA and HPV31, HPV16, HPV18, HPV52, HPV58, HPV33, HPV45, HPV6 or HPV11 pseudovirus neutralization experiments. Results: Nine HPV31 conformation-dependent neutralizing mAbs were screened, and three of which were HPV31 specific mAbs and the other six were cross-neutralizing mAbs.The mAbs whith IC₅₀<10 ng/mL were XM31-13 (0.36), XM31-19 (7.14), XM31-20 (6.9), XM31-23 (7.10). Conclusion: Nine conformation-dependent neutralizing mAbs were obtained, and four of them have low IC₅₀ value, which is useful for development of HPV31-VLP containing multivalemt vaccines.