乙肝核酸疫苗pCI/S2S接种小鼠后的免疫应答研究

目的 构建乙肝核酸疫苗pCI/S2S,研究其接种小鼠后,接种局部肌肉特异性抗原蛋白的表达以及所诱生的免疫应答反应.方法 从慢性乙肝患者的混合血清中制备HBV DNA模板,扩增出基因片段S2S,与pCI载体进行连接,得到重组质粒pCI/S2S.将其肌肉注射接种BALB/c小鼠,在不同的时间点检测血清中抗-HBs及抗-preS2;并于第12周时用细胞毒性检测试剂盒(LDH)检测体外细胞毒性T细胞(CTL)活性,同时以免疫组织化学法检测小鼠接种部位HBsAg的表达.结果 小鼠肌肉接种pCI/S2S完成后1周血清抗-HBs、抗-preS2开始阳转,阳转率于6周或8周达高峰,且能诱生特异性CTL应答.此外,免疫组织化学法能在小鼠的接种局部肌细胞中检测出目的抗原蛋白的表达.结论 乙肝核酸疫苗pCI/S2S接种小鼠后,能在小鼠体内诱生特异性的体液免疫和细胞免疫应答.

The Study about Immune Response to BALB/c Mice Inoculated with Hepatitis B Virus DNA Vaccine pCI/S2S

OBJECTIVE: To construct the hepatitis B virus (HBV) DNA vaccine pCI/S2S, and study the expression of specific antigen protein in muscle tissue section and the specific immune responses of humoral and cellular immunity in BALB/c mice being inoculated with pCI/S2S. METHODS: DNA templates were acquired from the mixed serum of patients with chronic hepatitis B, and the gene fragment S2S of HBV was cloned. The pCI vector and DNA S2S fragment were doubly cut by Kpn I and Not I, and then linked. HBV DNA vaccine pCI/S2S were intramuscularly injected into BALB/c mice, and boosted after 4 and 8 week. The expressing protein product in muscle tissue in situ and the specific cellular immune response were detected in 12 week after the first injection, the specific humoral immune responses were detected at 1, 2, 4, 6, 8, 12 week after the first injection. RESULTS: After pCI/S2S intramuscularly injected into BALB/c mice, HBsAb and preS2Ab were gradually determined with cytotoxic T lymphocyte responses induced, and HBsAg protein was expressed in situ in muscle tissue. CONCLUSION: pCI/S2S is effective HBV DNA vaccine that can induce the specific humoral and cellular immune responses.