Sun X, Si L, Cao Z, Wang Y, Liu T, Guo J
Institute of Immunopathology, Xi'an Medical University, Xi'an 710061, China.
Chin Med J (Engl). 2000 Mar;113(3):277-80.
To test the immunogenicity of recombinant plasmid DNA containing human papillomavirus type 16-L1 (HPV16-L1) coding sequence of mice.
The HPV16-L1 encoding sequence was generated by polymerase chain reaction (PCR), and inserted into TA cloning vector PCR II, then cloned in the eukaryotic expression vector pcDNA3.1 with CMV promoter. The recombinant plasmid DNA pcDNA-L1 was transferred into Cos-7 cells and used to immunize BALB/c mice via muscular injection. The expression of HPV16-L1 in transferred cells was identified by immunospot and immunocytochemistry, which tested specific anti-HPV16-L1 antibody in the serum of immunized mice.
Using the immunospot technique, we found L1 protein expression in pcDNA-L1 transferred cells. The immunocytochemistry studies demonstrated that the L1 protein was located in nuclei. In immunized mice, specific anti-HPV16-L1 antibodies could be detected by immunospot and immunocytochemistry 28 days after the first immunization and last at least 41 days.
We constructed HPV16-L1 eukaryotic expressing plasmid whose DNA could induce immunohumoral response in mice. This observation will be helpful in designing HPV16 prophylactic vaccine.
检测含人乳头瘤病毒16型L1（HPV16-L1）编码序列的重组质粒DNA对小鼠的免疫原性。
通过聚合酶链反应（PCR）产生HPV16-L1编码序列，将其插入TA克隆载体PCR II，然后克隆到带有巨细胞病毒（CMV）启动子的真核表达载体pcDNA3.1中。将重组质粒DNA pcDNA-L1转入Cos-7细胞，并通过肌肉注射用于免疫BALB/c小鼠。通过免疫斑点法和免疫细胞化学法鉴定转染细胞中HPV16-L1的表达，并检测免疫小鼠血清中的特异性抗HPV16-L1抗体。
使用免疫斑点技术，我们在pcDNA-L1转染细胞中发现了L1蛋白表达。免疫细胞化学研究表明L1蛋白位于细胞核中。在免疫小鼠中，首次免疫后28天通过免疫斑点法和免疫细胞化学法可检测到特异性抗HPV16-L1抗体，且至少持续41天。
我们构建了HPV16-L1真核表达质粒，其DNA可在小鼠中诱导免疫体液反应。这一观察结果将有助于设计HPV16预防性疫苗。
