中文摘要：

人乳头瘤病毒（human papillomavirus,HPV）58型是宫颈癌的主要诱因之一.HPV58在亚洲地区宫颈癌组织中的检出率仅次于HPV16/18.HPV58中和单克隆抗体可用于HPV病毒样颗粒（virus-like particle,VLP）疫苗的研究,并为病毒感染入侵机制的研究提供实验材料.本研究采用HPV58 L1 VLP免疫BALB/c小鼠,取其脾细胞进行杂交瘤细胞的制备,通过VLP-ELISA和假病毒中和实验筛选杂交瘤细胞株;经rProtein A纯化阳性杂交瘤细胞培养上清获得单抗;采用ELISA测定型别特异性中和单抗的亲和力,采用相加实验及变性VLP-ELISA分析单抗识别表位的性质;选取高亲和力单抗建立定量分析HPV58 L1 VLP的ELISA方法.获得了2株HPV58特异性中和单抗XM-22和XM-23,亲和常数分别为2.7×107mol-1.L和1.9×106mol-1.L,二者识别表位可能不同.同时获得2株具有交叉中和活性的单抗XM-21和XM-24,除可较高水平中和HPV58外,还可分别交叉中和亲缘关系较远的HPV18和HPV6.以XM-22建立的ELISA方法定量分析HPV58 L1 VLP的检测范围为0.05μg/mL～0.40μg/mL.本研究建立的ELISA方法可用于HPV58 L1 VLP疫苗生产的质量控制研究,获得的4株具有不同特点的中和单抗可用于HPV58感染入侵机制的研究.

英文摘要：

Persistent infections with human papillomavirus（HPV） 58 have been proved to be an important etiological factor of cervical cancer.Epidemiological study in Asia shows that HPV58 is a prevalent high-risk type,second only to HPV16/18.Neutralizing monoclonal antibodies（mAbs） against HPV58 can be used in HPV58 vaccine development and mechanism elucidation of virus invasion.Female BALB/c mice were immunized with HPV58 L1 virus-like particles（VLPs）.Hybridoma cells were prepared and neutralizing mAb secreting cell strains were identified by VLP-ELISA and pseudovirus neutralization assay.mAbs were purified by rProtein A chromatography from hybridoma cell culture supernatants.Affinities of type-specific neutralizing mAbs were analyzed by ELISA,and properties of mAbs were further identified by ELISA using denatured VLPs and additivity assay.Two type-specific neutralizing mAbs against HPV58,XM-22 and XM-23,were obtained,and affinity constants of them were 2.7×107 mol-1·L and 1.9×106 mol-1·L,respectively.The epitope recognized by XM-22 might be different from that recognized by XM-23.Two cross-neutralizing mAbs,XM-21 and XM-24,which could neutralize both HPV58 and heterogeneous HPVs such as HPV18/6,were also obtained.Concentrations of HPV58 L1 VLPs in samples ranged from 0.05 μg/mL to 0.40 μg/mL could be analyzed accurately by XM-22-based indirect ELISA.The established ELISA could be used in HPV58 L1 VLP vaccine production and four neutralizing mAbs with different characteristics would be helpful for mechanism study on HPV58 invasion.