Qing Rui, Xiang Qingke, Liu Zhongqi, Xiao Fei, Yang Fan
Department of Pathogenic Biology, School of Laboratory Medicine, Shaoyang University, Shaoyang 422600, China.
Department of Basic Medicine, Xiangnan University, Chenzhou 423000, China. *Corresponding author, E-mail:
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2018 Feb;34(2):123-128.
Objective To investigate the effect of aluminum phosphate (AP) and aluminum hydroxide (AH) as adjuvants on Brucella outer membrane protein 31 (Omp31) in inducing humoral and cellular immune responses and immune protection. Methods AP and AH adjuvants were prepared and separately mixed with Brucella Omp31 protein to measure the adsorption rates. The AP- and AH-absorbed Omp31 protein were intraperitoneally injected into BLAB/c mice at 0, 2, and 4 weeks, and meanwhile, unabsorbed Omp31 protein and PBS were used as controls. The levels of serum IgG, IgG1, IgG2a and genital tract secretion sIgA were determined by ELISA at 0, 2, 4 and 6 weeks. Spleen cells were collected for culture at 6 weeks, and the cells were stimulated by Omp31 for 48 hours followed by the analysis of IFN-γ and IL-10 levels in the supernatants by ELISA, and the determination of lymphocyte proliferation by CCK-8 assay. The mice were challenged with Brucella at 6 weeks, and bacterial content in spleen tissue was determined 1 and 2 weeks later. Results AP and AH could absorb over 70% and 85% of the Omp31 protein, respectively, for solutions at all the tested concentrations. ELISA suggested that serum IgG, IgG1, IgG2a and genital tract sIgA levels peaked 2 weeks after the last immunization for both AP and AH groups, and antibody level was higher in the AP and AH groups than the control groups, and higher in the AH group than in the AP group. CCK-8 assay showed that the proliferating rate of lymphocytes induced by the AH group was significantly higher than that by the AP group, and the AH group also showed significantly higher IFN-γ level in the supernatant than the AP group, but no significant difference in IL-10 level. The AH group had remarkably lower bacterial load in the spleen than the AP group 2 weeks after challenged by Brucella 16M strain. Conclusion Both AP and AH adjuvants effectively enhanced immunogenicity and immune protection of the Brucella Omp31 protein, and AH was superior to AP in this respect.
目的 探讨磷酸铝(AP)和氢氧化铝(AH)作为佐剂对布鲁氏菌外膜蛋白31(Omp31)诱导体液免疫和细胞免疫应答及免疫保护的影响。方法 制备AP和AH佐剂,并分别与布鲁氏菌Omp31蛋白混合,测定吸附率。将吸附有AP和AH的Omp31蛋白分别于0、2、4周腹腔注射到BLAB/c小鼠体内,同时,将未吸附的Omp31蛋白和PBS作为对照。在0、2、4和6周时通过ELISA测定血清IgG、IgG1、IgG2a和生殖道分泌物sIgA水平。在6周时收集脾细胞进行培养,用Omp31刺激细胞48小时,然后通过ELISA分析上清液中IFN-γ和IL-10水平,并通过CCK-8法测定淋巴细胞增殖。在6周时用布鲁氏菌攻击小鼠,1和2周后测定脾组织中的细菌含量。结果 对于所有测试浓度的溶液,AP和AH分别可吸附70%以上和85%以上的Omp31蛋白。ELISA结果表明,AP和AH组血清IgG、IgG1、IgG2a和生殖道sIgA水平在末次免疫后2周达到峰值,且AP和AH组抗体水平高于对照组,AH组高于AP组。CCK-8法显示,AH组诱导的淋巴细胞增殖率显著高于AP组,AH组上清液中IFN-γ水平也显著高于AP组,但IL-10水平无显著差异。用布鲁氏菌16M菌株攻击后2周,AH组脾内细菌载量明显低于AP组。结论 AP和AH佐剂均能有效增强布鲁氏菌Omp31蛋白的免疫原性和免疫保护作用,且在这方面AH优于AP。