Objective To study the hereditary stability of master seed bank (MSB) and working seed bank (WSB) of recombinant adenoviral vector which express human papillomavirus type 16 E6E7 fusion protein (AdHPV16E6E7).Methods The infectivity titers of Ad-HPV16E6E7 MSB and WSB were evaluated by microscope check and virus infectious titer assay.The stability of HPV16E6E7 gene was evaluated by PCR amplification.The protein expression of Ad-HPV16E6E7 was evaluated by Western blot and immunofluorescence.The biological effect of AdHPV16E6E7 was evaluated by tumor inhibition assay using C57BL/6 mice which inoculated with TC-1 tumor cells.Results The infectivity titers of MSB and WSB of Ad-HPV16E6E7 were 6.31 × 109 IU/mL and 3.00 × 109 IU/mL,respectively.HPV16E6E7 gene and protein were stable.When the infectivity titers of MSB and WSB were 109 IU/mL,the antitumor rate was 100％ in mouse model and showed statistical significance compared to control group (x2 =20.00,20.00,P all ＜ 0.01).Conclusions The hereditary stability of MSB and WSB of Ad-HPV16E6E7 are stable,which demonstrate that the vector seeds meet the requirements of vaccine development in the future.%目的 了解表达人乳头瘤病毒(HPV) 16型E6和E7融合蛋白的重组腺病毒载体(Ad-HPV16E6E7)疫苗的主种子库(MSB)和工作种子库(WSB)的遗传稳定性.方法 通过显微镜检查和病毒感染效价测定实验测定Ad-HPV16E6E7 MSB和WSB的感染性滴度;采用PCR扩增法鉴定HPV16E6E7目的基因稳定性;采用Western印迹法和免疫荧光法鉴定Ad-HPV16E6E7目的蛋白表达;采用C57BL/6小鼠TC-1肿瘤细胞生长抑制试验评价Ad-HPV16E6E7的生物学效应.结果 Ad-HPV16E6E7的MSB和WSB的感染性滴度分别为6.31×109 IU/mL和3.00×109 IU/mL;插入目的基因和表达目的蛋白稳定;小鼠TC-1肿瘤生长抑制试验表明MSB和WSB滴度的数量级在109 IU/mL时,小鼠抑瘤率达100％,与对照组相比差异均有统计学意义(x2=20.00、20.00,P均＜0.01).结论 Ad-HPV16E6E7疫苗的种子库遗传稳定,符合将来疫苗研发的要求.
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