摘要目的 探讨二甲基三十六烷基铵(DDA)、卡介苗多糖核酸(BCG-PSN)作为结核病融合蛋白疫苗佐剂的免疫效应.方法 采用热酚法制备BCG-PSN,与DDA联合作为融合蛋白AMM(Ag85B-MPT64190-198-Mth8/4)佐剂免疫小鼠,设卡介苗、生理盐水、弗氏不完全佐剂(IFA)及单独应用DDA或BCG-PSN一种佐剂的各组作为对照.应用ELISA和酶联免疫斑点法检测免疫小鼠的体液与细胞免疫反应.结果 分离培养疫苗免疫小鼠的脾脏淋巴细胞,用特异性抗原AgSSB刺激后,AMM+DDA+BCG-PSN疫苗组、AMM+DDA组和卡介苗组小鼠每1×106个脾淋巴细胞分泌γ-干扰素的细胞数分别为222±79、259±85和230±64,均明显高于AMM+PBS组(40±4)、AMM+IFA组(10 ±3)、AMM+BCG-PSN组(132±18)和生理盐水组(8±4),差异有统计学意义(t值为2.923～7.118,均P<0.05);与单用DDA组比较,添加DDA和BCG-PSN两种佐剂的疫苗组小鼠血清抗体滴度Ig2a/IgG1较高(0.125和0.025),但γ-干扰素分泌水平没有增高.结论 AMM+DDA+BCG-PSN结核病亚单位疫苗能够引起较强的Th1细胞免疫为主的免疫反应,DDA+BCG-PSN(尤其是DDA)具有增强结核病亚单位疫苗Th1细胞免疫应答的佐剂效应.

abstractsObjective To investigate the activity of a novel adjuvant consisting of dimethyl-dioctyldecyl ammonium bromide (DDA) and BCG polysaccharide nucleic acid (BCG-PSN). Methods BCG-PSN was extracted by hot-phenol method, and combined with DDA and Mycobacterium tuberculosis fusion antigen AMM (Ag85B-MPT64190-198-Mtb8.4) to formulate the Mycobacterium tuberculosis subunit vaccine. Mice were iannunized subcutaneously with a 2-week interval between the immunizations (0. 2 ml/ dose), and humoral and cell-mediated immunity were detected by ELISA and ELISPOT respectively. Results With the stimulation of Ag85B in vitro, the number of antigen specific IFN-γ producing spleen lymphocytes were 222 ± 79,259 ± 85,230 ± 64 per million respectively in the mice immunized with AMM + DDA + BCG-PSN, AMM + DDA, and BCG. Spleen lymphocytes in these 3 groups produced higher levels of IFN-γ compared to the groups with the adjuvant of IFA or BCG-PSN alone or without adjuvant upon stimulation with Ag85B (t = 2.923 - 7. 118, P < 0.05). Furthermore, the adjuvant consisting of DDA and BCG-PSN increased the ratio of Ig2a/IgG1 than DDA alone (0. 125 vs 0. 025). Combined with AMM, the adjuvant DDA and the one consisting of DDA and BCG-PSN induced higher level of immunity than incomplete Freund' s adjuvant (IFA), NaCl, and BCG-PSN alone. Conclusion Mycobacterium tuberculosis subunit vaccine AMM + DDA + BCG-PSN induced a strong Thl-type immune response, and DDA + BCG-PSN, especially DDA promoted the immune response of the M. tuberculosis subunit vaccine in mice.