【摘要】 目的对HPV18型DNA疫苗的免疫原性和安全性进行初步评价。方法将HPV18L1基因与真核表达载体pcDNA3.1(+)连接,构建DNA疫苗质粒pcDNAL1。使用脂质体将重组质粒转染COS7细胞,用Westernblot检测体外瞬时表达的抗原。将重组质粒pcDNAL1肌肉注射BALBc小鼠(100μg只),加强免疫3次后分别采集血清,用ELISA法和Westernblot法检测抗体,实时荧光定量PCR法检测质粒的组织分布。采用ELISA法检测DNA疫苗免疫后抗核抗体和抗双链DNA抗体的效价。结果重组质粒可在真核细胞中有效地转录和表达HPV18L1蛋白。小鼠免疫后可检出较高滴度的抗HPV18L1抗体和特异条带。免疫后质粒主要分布在注射部位,且随着时间延长被清除;未检测到抗核抗体和抗双链DNA抗体。结论HPV18L1重组质粒可诱导小鼠产生特异的体液免疫反应,且该DNA疫苗是安全的,为进一步研制开发HPVDNA疫苗打下了基础。更多还原

【Abstract】 Objective To primarily evaluate the immunogenicity and safety of HPV18 L1 DNA vaccine.Methods Insert HPV18 L1 gene into eukaryotic expression vector pcDNA3.1(+) to construct recombinant plasmid pcDNA-L1 as a DNA vaccine.Transfect COS-7 cells with the recombinant plasmid in the mediation of liposome,and identify the antigen transiently expressed in vitro by Western blot.Inject ~BALB/c mice with recombinant plasmid pcDNA-L1 for 4 times,each at a dosage of 100 μg.Collect the sera of mice for detection of antibody by ELISA and Western blot.The distribution of DNA in tissue was analyzed by real-time fluorescent quantitative PCR.The titers of anti-nuclear and anti-dsDNA antibodies were detected by ELISA.Results The recombinant plasmid pcDNA-L1 was effectively transcribed in eukaryotic cells and HPV18 L1 protein was successfully expressed.Specific anti-HPV18 L1 antibodies at high titers were detected in the sera of immunized mice.After immunization,the recombinant plasmid pcDNA-L1 was mainly distributed in the injection site and was removed as the time goes on.Neither anti-nuclear nor anti-dsDNA antibody was detected.Conclusion HPV18 L1 as a DNA vaccine was safe and induced specific humoral immune response in mice.It laid a foundation of further development of HPV DNA vaccine.更多还原