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目的 建立稳定表达核蛋白(NP)的P815细胞株,并以此作为CTL的靶细胞,研究汉滩病毒(HTHV)诱导产生的NP抗原特异性CTL。方法 脂质体介导pEFBOS/HTHVNP和pDORneo质粒DNA共转染P815细胞株,用免疫组化染色法证实NP在胞浆中的表达。分别取HTHV感染小鼠的脾细胞和pEFBOS/HTHVNP质粒DNA免疫小鼠的脾细胞,经体外抗原和IL2再刺激诱导产生CTL效应细胞,进行4h51Cr释放实验。结果 杀伤试验结果表明,无论通过病毒感染还是基因免疫获得的CTL效应细胞,均可以特异性杀伤NP基因转染的靶细胞。结论 HTHVNP可以有效诱导CTL的产生,该研究结果为进一步设计HTHV合成肽疫苗或基因疫苗提供了重要实验依据。
Objective To establish P815 cell line stably expressing nucleoprotein (NP) and use it as the target cell of CTL to study the NP antigen-specific CTL induced by Hantaan virus (HTHV). Methods P815 cells were co-transfected with plasmid pEFBOS / HTHVNP and pDORneo by liposome. The expression of NP in cytoplasm was confirmed by immunohistochemical staining. The spleen cells of mice immunized with HTHV-infected mice and pEFBOS / HTHVNP plasmid DNA were respectively immunized with CTL effector cells induced by restimulation with IL-2 in vitro and 4h51Cr release assay. Results The results of killing test showed that CTL effector cells obtained by virus infection or gene immunization could specifically kill target cells transfected with NP gene. Conclusion HTHVNP can effectively induce the production of CTL. The results of this study provide important experimental evidence for the further design of HTHV synthetic peptide vaccine or gene vaccine.