摘要
目的研究弓形虫棒状体蛋白2(ROP2)和膜表面蛋白1(SAG1)重组复合抗原(ROP2-SAG1)的核酸和蛋白两种疫苗的免疫效应。方法 60只雌性BALB/c小鼠随机均分为4组(每组15只),即rROP2-SAG1蛋白组、无菌生理盐水组、pcROP2-SAG1质粒组和pcDNA3.1质粒组,rROP2-SAG1组以2.5μgrROP2-SAG1重组蛋白与等体积佐剂乳化后,背部皮下多点免疫小鼠;无菌生理盐水组以等体积无菌生理盐水替代重组蛋白,首次免疫使用福氏完全佐剂,其余使用福氏不完全佐剂。pcROP2-SAG1组和pcDNA3.1组分别以pcROP2-SAG1和空质粒pcDNA3.1腿部肌肉注射免疫,剂量为100μg/次;以上各组小鼠共免疫3次,每次间隔2周。采用间接ELISA方法检测免疫后25、45和70d小鼠血清中特异性IgG抗体,及末次免疫后2周小鼠血清特异性IgG1和IgG2a抗体。用CCK-8细胞增殖法检测小鼠脾细胞增殖,间接ELISA法检测小鼠脾细胞培养上清液中γ干扰素(IFN-γ)和白介素2(IL-2)表达水平。结果 rROP2-SAG1组小鼠血清特异性IgG抗体水平随着免疫时间的延长持续升高,pcROP2-SAG1质粒组抗体则随着免疫时间的延长相对稳定。末次免疫后2周,rROP2-SAG1组小鼠血清IgG1抗体水平(1.538±0.183)显著高于IgG2a(0.618±0.122)(P<0.05),而pcROP2-SAG1组IgG1抗体水平(1.107±0.137)与IgG2a(0.830±0.185)间的差异无统计学意义(P>0.05)。以特异性抗原rROP2-SAG1为刺激物时,rROP2-SAG1组小鼠脾细胞受特异性抗原刺激的增殖效果(A450值=0.348±0.042)显著高于pcROP2-SAG1组(0.123±0.018)(P<0.05)。rROP2-SAG1组小鼠脾细胞培养上清液中IFN-γ含量[(149.37±30.51)pg/ml]、IL-2含量[38.58±9.10)pg/ml],分别与pcROP2-SAG1组IFN-γ[(138.58l±29.92)pg/ml]、IL-2[37.47l±9.26)pg/ml]比较,差异均无统计学意义(P>0.05)。结论 ROP2-SAG1重组复合蛋白诱导的抗体水平、脾细胞增值效应显著高于重组质粒pcROP2-SAG1。
Objective To study the immune response elicited by the recombinant protein vaccine and DNA vaccine of the complex antigen ROP2-SAG1 from Toxoplasma gondii. Methods Sixty female BALB/c mice were randomly divided into 4 groups （15 per group）. Mice in rROP2-SAG1 group were immunized subcutaneously with 2.5 μg rROP2-SAG1 protein formulated in Freund′s adjuvant. Mice in control group received only adjuvant emulsified with normal saline. Mice in recombinant plasmid pcROP2-SAG1 and control plasmid pcDNA3.1 groups were each injected intramuscularly with 100 μg of pcROP2-SAG1 and pcDNA3.1, respectively. All the mice received three immunizations at 2-week intervals. Serum samples were collected at 25, 45, and 70 days after immunization for determining antibody IgG, and at 2 weeks after the last immunization IgG1 and IgG2a were detected all by ELISA. Cell counting kit-8 （CCK-8） was used to deter- mine the splenocyte proliferation, and the supernatant of cultured splenocytes was collected for the detection of IFN-γ by ELISA. Results The level of IgG continued to rise in rROP2-SAG1 group after immunization, and similarly in pcROP2- SAG1 group. At 2 weeks after the last immunization, level of IgG1 （1.538±0.183） was higher than that of IgG2a （0.618± 0.122）（P0.05） in rROP2-SAG1 group. Whereas no significant difference between IgG1 （1.107±0.137） and IgG2a （0.830± 0.185） was observed in pcROP2-SAG1 group （P0.05）. Compared with the pcROP2-SAG1 group （A450=0.123±0.018）, more significant proliferation response of splenocytes was observed in rROP2-SAG1 group （0.348±0.042） （P0.05）. There was no significant difference（P0.05） of IFN-γ and IL-2 in the supernatant of cultured splenocytes between the groups of rROP2- SAG1 and pcROP2-SAG1. Conclusion The antibody level and splenocyte proliferation have been significantly higher in mice immunized with recombinant protein rROP2-SAG1 than those with recombinant plasmid pcROP2-SAG1.
出处
《中国寄生虫学与寄生虫病杂志》
CAS
CSCD
北大核心
2010年第5期359-363,共5页
Chinese Journal of Parasitology and Parasitic Diseases
基金
浙江省自然科学基金(NoY205567)
浙江省医药卫生科学研究基金(No2007B137)~~
