恶性疟原虫富组氨酸蛋白-2基因诱导的免疫应答研究

目的 ]探讨编码恶性疟原虫富组氨酸蛋白 2 (HRP Ⅱ )C端基因的真核表达重组质粒诱导小鼠的体液和细胞免疫效果。 方法 ]将起始码和终止码引入HRP ⅡC端基因片段两端 ,经测序鉴定读框 ,将包含起始码和终止码的HRP Ⅱ基因片段克隆入真核表达质粒pcDNA3 1( )中 ,进行酶切鉴定。用重组真核表达质粒pcDNA3 1( ) /HRP Ⅱ 经肌肉免疫小鼠 3次 ,每次间隔 3wk。第 3次免疫后 2wk ,取小鼠血清和脾细胞 ,分别用ELISA测定HRP Ⅱ抗体水平和用脾细胞增殖实验测定细胞免疫反应。 结果 ]序列测定结果表明 ,HRP ⅡC端基因片段被准确地引入起始码和终止码 ;酶切鉴定表明包含起始码和终止码的HRP ⅡC端基因片段成功地克隆入pcDNA3 1( ) ,形成pcDNA3 1( ) /HRP Ⅱ 。在pcDNA3 1( ) /HRP Ⅱ 免疫鼠血清中可检测出高水平的HRP Ⅱ抗体 ,用原核表达的HRP Ⅱ蛋白刺激免疫脾细胞 ,可检测出明显的细胞增殖反应。 结论 ]编码HRP Ⅱ的真核表达重组质粒可诱导小鼠产生明显的体液和细胞免疫反应。其有可能作为恶性疟原虫红内期复合DNA疫苗的候选基因

IMMUNE RESPONSE IN MICE INDUCED BY C TERMINAL ENCODING GENE OF PLASMODIUM FALCIPARUM HISTIDINE RICH PROTEIN 2

OBJECTIVE: To explore the humoral and cellular immune responses in mice to eukaryotic expression recombinant plasmid encoding histidine rich protein 2 (HRP-II) of Plasmodium falciparum. METHODS: The start and stop codes were introduced into HRP-II gene fragment, the reading frame and the position of start and stop codes in HRP-II were identified by sequencing. HRP-II fragment containing the start and stop codes was cloned into pcDNA3.1 (-) to form pcDNA3.1 (-)/HRP-II. The BALB/c mice were immunized i.m. with the plasmids for 3 times in 3 weeks intervals. Two weeks after the last immunization, the sera and splenocytes were collected to investigate anti-HRP-II antibodies by ELISA and the splenocytes proliferation response to HRP-II. RESULTS: Sequence data show that the reading frame and the position of start and stop codes are correct. Restriction enzyme digestion indicated that the HRP-II gene fragment containing start and stop codes was successfully cloned into pcDNA3.1 (-). Mice raised significant anti-HRP-II antibodies after pcDNA3.1 (-)/HRP-II immunization, and the splenocytes proliferated prominently when stimulated with HRP-II protein. CONCLUSION: Eukaryotic expression recombinant plasmid encoding HRP-II gene can induce significantly humoral and cellular immune response in mice. HRP-II gene may be a good candidate for P. falciparum blood-stage multiple DNA vaccine.