摘要
目的观察毕赤酵母菌表达弓形虫GRA1蛋白免疫效果。方法采用皮下注射法免疫接种小鼠,末次免疫后第2周用ELISA检测其特异性抗体水平;流式检测鼠脾淋巴细胞亚群,末次免疫后第3周,用弓形虫RH株速殖子分别攻击感染疫苗免疫组和对照组,观察小鼠感染情况及存活时间。结果ELISA结果表明,纯化GS115/pPIC9K-GRA1表达的蛋白和GS115/pPIC9K-GRA1表达的混合蛋白组抗体水平高于佐剂组、NS对照组及GS115/pPIC9K表达的混合蛋白组(P<0.05);流式结果显示,各组CD4+淋巴细胞细胞数量变化无显著性差异(P>0.05),而CD8+淋巴细胞数量则明显增多,纯化GS115/pPIC9K-GRA1表达的蛋白、GS115/pPIC9K-GRA1表达的混合蛋白组、GS115/pPIC9K表达的混合蛋白组及佐剂组明显高于NS组(P<0.05);小鼠攻击实验表明,与NS及GS115/pPIC9K表达的混合蛋白组相比,纯化GS115/pPIC9K-GRA1表达的蛋白和GS115/pPIC9K-GRA1表达的混合蛋白组小鼠存活时间显著延长(P<0.05)。结论毕赤酵母菌表达弓形虫GRA1蛋白免疫小鼠后可诱导特异性抗体产生,CD4+淋巴细胞增殖不明显,而CD8+淋巴细胞则显著增殖及对弓形虫攻击感染有一定的保护作用。
Objective To observe the specific immune responses and immunity protection of protein of GRA1 expressed in Pichia pastoris. Methods After preparation and purification of protein , mice were immunized subcutaneously with GRA1. Two weeks after the last immunization, sera were respectively collected from each group. Leukomonocyte subpopulation was analyzed with BD Flow cytometer, specific antibody levels were measured by ELISA. The mice in immunized groups and control groups were challenged with T. gondii tachyzoites, and then the survival time was compared. Results Two weeks af- ter the last immunization, ELISA showed levels of specific antibody in sera of mice inoculated with purification GRA1 expressed in GS115/pPIC9K - GRA1 and admixture protein expressed in GS115/pPIC9K - GRA1 were significantly higher than admixture protein expressed in GSllS/pPIC9K and those in control group mice. The number of CD4 ^＋ T cell did not increased apparently compared with each groups （P 〉 0.05）. However, CD8^＋ T cell was sigificant increased than those in NS control group （P 〈 0.01 ） and the value of CD4 ^＋/CD8^＋ was decreased. After challenging with violent virulence strain of tachyzoites, the mean survival time of immunized mice groups was significantly longer than that in control groups and admixture protein ex- pressed in GS115/pPIC9K. Conclusion It can induce specific antibody response and CD8^＋ T cell was sigificant increase than those in NS control group and partial protection against challenge of virulence strain of tachyzoites which expressed GRA1 in Pichia pastoris.
出处
《中国热带医学》
CAS
2008年第9期1485-1487,共3页
China Tropical Medicine
基金
湖南省卫生厅资助项目(NoC2006 -033)
湖南环境生物职业技术学院(NoZ07 -09)
关键词
弓形虫
GRAl
毕赤酵母菌
接种
免疫
Toxoplasma gondii
GRA1
Pichia pastoris
Protein vaccination
Immunity effect
