摘要
目的 观察乙型肝炎病毒表面抗原 (HBsAg)基因表达质粒 (pS)及其与鼠白细胞介素 18基因融合表达质粒 (p18S)的构建及其诱导BALB/c鼠免疫应答的能力。 方法 用聚合酶链反应(PCR)法获得的S基因克隆到真核表达质粒 pcDNA3.1+上 ,构建 pS。另用三步PCR法将小鼠白细胞介素 18基因融合到S基因的 5′端 ,再插入到 pcDNA3.1+的EcoRI位点上 ,构建了 p18S。分别免疫 10只和 6只BALB/c小鼠 ,阴性对照用 pcDNA3.1+免疫 4只BALB/c小鼠。用酶联免疫方法检测每份血清的抗 HBs的效价。采用3 H标记法检测脾细胞HBsAg特异性细胞毒反应。 结果 pS质粒免疫BALB/c小鼠多数于 4周时出现抗 HBs,最高可达 5 30mIU/ml,平均为 135mIU/ml。p18S质粒也能诱导低水平的抗 HBs ,平均为 2 0mIU/ml。pS和 p18S诱导的HBsAg特异性的细胞毒活性分别为 37.1%和 34 % ,而 pcDNA3.1+载体质粒免疫小鼠血清未能检出抗 HBs,细胞毒活性为 13.2 %。结论 pS质粒能够有效地诱导BALB/c小鼠的体液和细胞免疫反应 ,白细胞介素 18基因与S基因融合表达质粒免疫似乎不能增强免疫应答 。
Objective To observe the specific humoral and cellular immune response in BALB/c mice injected with pS and p18S. Methods pS and p18S were constructed separately by inserting HBsAg gene fragment and the fusion gene fragment of HBsAg and mouse interleukin 18(IL 18) into the reading frame of pcDNA3.1+. Mice were injected with either plasmid intramuscularly in a total dose of 300 μg per mouse. Every serum sample was detected for anti HBs using enzyme linked immunosorbent assay(ELISA). Furthermore, HBsAg specific cytotoxic T lymphocytes activity was measured. Results The expression of HBsAg was demonstrated by ELISA in p815 cells transfected with pS and p18S. pS can stimulate a positive antibody response. The average level was 135 mIU/ml, with the highest level of 530 mIU/ml. p18S could elicit relatively lower antibody response which was 20 mIU/ml. HBsAg specific CTL activities were 37.1% and 34% separately in pS and p18S immunized mice. It was only 13.2% when detected in pcDNA3.1+ immunization. Conclusion pS is effective to stimulate a humoral and cellular response in H 2d mice. IL 18 gene can not enhance the immune response when fused with HBsAg gene. Conversely, it seems to inhibit an immune response.
出处
《中华传染病杂志》
CAS
CSCD
北大核心
2001年第2期77-80,共4页
Chinese Journal of Infectious Diseases
